Transposon mutagenesis of nuclear photosynthetic genes in Zea mays

The discovery of a new maize (Zea mays L.) transposon system, Mutator, and the cloning of the 1.4 kilobase transposon, Mul, have made feasible the isolation of nuclear photosynthetic genes which are recognized only by their mutant phenotype. Mutant maize plants which express a high chlorophyll fluorescent (hcf) phenotype due to a defect in the electron transport or photophosphorylation apparatus have been isolated following mutagenesis with an active Mutator stock. The affected genes and their products in these mutants are inaccessible to classical methods of analysis. However, mutagenesis with the Mutator transposon makes it possible to isolate these genes.Although the PSII-deficient mutant hcf3 has been thoroughly studied by classical photo-biological methods, the nature of the lesion which results in the observed phenotype has not been established. A Mutator-induced allele of hcf3 has been isolated. A fragment of genomic DNA has been identified which is homologous to Mul and co-segregates with the mutant phenotype. This fragment is expected to contain a portion of the hcf3 locus which will be used to clone the normal gene. Direct study of the gene can provide insight into the nature and function of its polypeptide product.This approach can be used to study any photosynthetic gene which has been interrupted by a transposon. The isolation of more than 100 different chemically-induced hcf mutants, most of which can not be fully characterized using classical means, indicates the wealth of information which can be obtained using a transposon tagging technique.     

玉米(Zea mays L.)叶脉发育的研究

玉米的叶脉在单子叶植物中有一定的代表性,叶脉由四级组成,粗细不同的一、二、三级叶脉均从叶基向叶尖方向延伸,属叶片的纵向叶脉,四级脉横向与一、二、三级叶脉连接,是横向的叶脉,各级叶脉有各自的形成方式,由于它们有规律的分布,从而构成了叶片的输导网络,各级叶脉的发生和发育与叫片的生长有直接的关系。     

A mutation causing proline requirement in Zea mays

Summary A conditional seedling lethal, monogenic recessive, endosperm mutant is described. Phenotypic can be accomplished when embryos are cultured in vitro on media supplemented with proline. The efficiency of the repair is proportional to the concentration of proline in the medium. Normal growth is resumed at a dose of 160 mg/l. All the data collected are most easily interpreted by assuming that the mutant, symbolized pro has a genetic block in the biosynthetic route leading to proline.This is probably the first case of a strict genetic requirement for an amino acid to be reported in Zea mays. The possible reasons for the difficulties encountered in isolating obligate auxotrophs in higher plants are briefly reviewed.     

Aluminium modulation of the photosynthetic carbon reduction cycle in Zea mays

Two-weeks-old maize (Zea mays L. cv. XL-72.3) plants were submitted to Al concentrations of 0-81 g m-3 for 20 d, after which the A1 concentration-dependent effects on CO2 uptake by the mesophyll tissue and subsequent CO2 assimilation in the photosynthetic carbon reduction cycle of bundle sheath cells were investigated. The net photosynthetic rate (PN) and stomatal conductance (gs) increased continuously up to 27 g m-3 Al, whereas the intercellular CO2 concentration showed minimum values with the 27 g m-3 Al treatment. Moreover, the starch and saccharide concentrations, and fructose-1,6-bisphosphatase did not change significantly with increasing Al concentrations. The photosynthetic electron transport rates along with photosystems 2 and 1 started falling from 9 g m-3 Al onwards, while thylakoid acyl lipid composition did not show a clear pattern. With the Al concentration at 81 g m-3, NADP-malate dehydrogenase activity decreased to minimum values, whereas the opposite occurred with those of pyruvate dikinase, NADP-malic enzyme, and phosphoenolpyruvate carboxylase. Thus in vivo Al concentrations modulate the photosynthetic reduction cycle, possibly by interacting with the carbon flow rate exported to the cytosol. Although the inhibition of NADP-malate dehydrogenase activity might limit pyruvate dikinase, NADP-malic enzyme, and phosphoenolpyruvate carboxylase activities, in vivo the balance between phosphoenolpyruvate production and its carboxylation remains unaffected.     

Aluminium modulation of the photosynthetic carbon reduction cycle in Zea mays

Two-weeks-old maize (Zea mays L. cv. XL-72.3) plants were submitted to Al concentrations of 0-81 g m-3 for 20 d, after which the A1 concentration-dependent effects on CO2 uptake by the mesophyll tissue and subsequent CO2 assimilation in the photosynthetic carbon reduction cycle of bundle sheath cells were investigated. The net photosynthetic rate (PN) and stomatal conductance (gs) increased continuously up to 27 g m-3 Al, whereas the intercellular CO2 concentration showed minimum values with the 27 g m-3 Al treatment. Moreover, the starch and saccharide concentrations, and fructose-1,6-bisphosphatase did not change significantly with increasing Al concentrations. The photosynthetic electron transport rates along with photosystems 2 and 1 started falling from 9 g m-3 Al onwards, while thylakoid acyl lipid composition did not show a clear pattern. With the Al concentration at 81 g m-3, NADP-malate dehydrogenase activity decreased to minimum values, whereas the opposite occurred with those of pyruvate dikinase, NADP-malic enzyme, and phosphoenolpyruvate carboxylase. Thus in vivo Al concentrations modulate the photosynthetic reduction cycle, possibly by interacting with the carbon flow rate exported to the cytosol. Although the inhibition of NADP-malate dehydrogenase activity might limit pyruvate dikinase, NADP-malic enzyme, and phosphoenolpyruvate carboxylase activities, in vivo the balance between phosphoenolpyruvate production and its carboxylation remains unaffected.     

The effects of development at sub-optimal growth temperatures on photosynthetic capacity and susceptibility to chilling-dependent photoinhibition in Zea mays

When plants of Zea mays L. cv. LG11 that have been grown at optimal temperatures are transferred to chilling temperatures (0–12°C) photoinhibition of photosynthetic CO₂ assimilation can occur. This study examines how growth at sub-optimal temperatures alters both photosynthetic capacity and resistance to chilling-dependent photoinhibition. Plants of Z. mays cv. LG11 were grown in controlled environments at 14, 17, 20 and 25°C. As a measure of the capacity for photosynthesis under light limiting conditions, the maximum quantum yields of CO₂ assimilation (φ^a.c) and O₂ evolution (φ^a.o) were determined for the laminae of the second leaves at photon fluxes of 50–150 μmol m^-2s^-1. To determine photosynthetic capacity at photon fluxes approaching light saturation, rates of CO₂ uptake (A₁₅₀₀) and O₂ evolution (A₁₅₀₀) were determined in a photon flux of 1500 μmol m^-2s^-1. In leaves developed at 14°C, φ and φ were 26 and 43%, respectively, of the values for leaves grown at 25°C. Leaves grown at 17°C showed intermediate reductions in φ and φ, whilst leaves developed at 20°C showed no significant differences from those grown at 25°C. Similar patterns of decrease were observed for A₁₅₀₀ and A_1500.0 with decreasing growth temperature. Leaves developed at 25°C showed higher rates of CO₂ assimilation at all light levels and measurement temperatures in comparison to leaves developed at 17 and 14°C. A greater reduction in A₁₅₀₀ relative to A_1500.0 with decreasing growth temperature was attributed to increased stomatal limitation. Exposure of leaves to 800–1000 μmol m^-2 s^-1 when plant temperature was depressed to ca 6.5°C produced a photoinhibition of photosynthetic CO₂ assimilation in all leaves. However, in leaves developed at 17°C the decrease in A₁₅₀₀ following this chilling treatment was only 25% compared to 90% in leaves developed at 25°C. Recovery following chilling was completed earlier in leaves developed at 17°C. The results suggest that growth at sub-optimal temperatures induces increased tolerance to exposure to high light at chilling temperatures. This is offset by the large loss in photosynthetic capacity imposed by leaf development at sub-optimal temperatures.     

Metabolism of acetate-14C in normal and opaque-2 Zea mays endosperm during development

Acetate-[2-¹⁴C]metabolism by developing normal and opaque-2 maize endosperms showed considerable differences in incorporation of label into organic     

The filifolium1 mutation perturbs shoot architecture in Zea mays (Poaceae)

Plant architecture is elaborated through the activity of shoot apical meristems (SAMs), which produce repeating units known as phytomers, that are comprised of leaf, node, internode, and axillary bud. Insight into how SAMs function and how individual phytomer components are related to each other can been obtained through characterization of recessive mutants with perturbed shoot development. In this study, we characterized a new mutant to further understand mechanisms underlying shoot development in maize. The filifolium1-0 (ffm1-0) mutants develop narrow leaves on dwarfed shoots. Shoot growth often terminates at the seedling stage from depletion of the SAM, but if plants survive to maturity they are invariably bushy. KN1-like homeobox (KNOX) proteins are inappropriately regulated in mutant apices, adaxial identity is not specified in mutant leaves, and axillary meristems develop precociously. We propose that FFM1 acts to demarcate zones within the SAM so that appropriate fates can be conferred on cells within those zones by other factors. On the basis of the mutant phenotype, we also speculate about different relationships between phytomer components in maize and Arabidopsis.     

Seed development and vivipary in Zea mays L.

Seed development was investigated in kernels of developing wild-type and viviparous (vp-1) Zea mays L. Embryos and endosperm of wild-type kernels began to dehydrate at approx. 35 d after pollination (DAP); viviparous embryos did not desiccate but accumulated fresh weight via coleoptile growth in the caryopses. Concentrations of endogenous abscisic acid (ABA) in the embryo were relatively high early in development, being approx. 150 ng·g^-1 fresh weight at 20 DAP. The ABA content declined thereafter, falling to approx. 50 ng·g^-1 at 30 DAP. Endosperm ABA content was always low, being less than 20 ng·g^-1. There were no differences between wild-type and vp-1 tissues. Immature kernels did not germinate when removed from the ear until late in development. The ability to germinate was correlated with decreasing moisture content in the endosperm at the time of removal; premature drying of immature kernels resulted in greatly increased germination following imbibition. Excised embryos germinated precociously when removed from the endosperm as early as 25 DAP. Such germination could be prevented by treatment with 10^-5 M ABA or by lowering the solute potential (_s) of the medium with 0.3 M mannitol. Treatment of excised embryos with ABA led to internal ABA concentrations comparable to those in embryos in which germination was inhibited in situ. Mannitol treatment did not have this effect, although water-deficit stress of excised embryos resulted in substantial ABA production. Germinated vp-1 embryos were less sensitive to growth inhibition by ABA or mannitol than germinating wild-type embryos. The vp-1 seedlings were not wilty and their transpiration rates were reduced in response to ABA or water shortage.Abbreviations and symbols ABA abscisic acid - DAP days after pollination - FW fresh weight - vp-1 viviparous genotype - _s solute potential     

Benzyladenine effects on the development of the photosynthetic apparatus in Zea mays: Studies on photosynthetic activity, enzymes and (etio)chloroplast ultrastructure

Primary leaf segments from 8-day-old dark-grown, and from 4- and 8-day-old light-grown seedlings of Zea mays L. cv. Fronica, were treated with 10^-bM benzyladenine (BA) in the dark for 14 h. The segments were then studied after an exposure to light for 14 h. Photosynthetic activity (O₂ evolution and CO₂ fixation) and chlorophyll accumulation were stimulated by BA in dark-grown leaf segments with etioplastids in the earliest stage of development. In these segments BA stimulated the activities of ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39), phosphoenolpyruvate carboxylase (EC 4.1.1.31), NADP⁺-malic enzyme (EC 1.1.1.40) and pyruvate, orthophosphate dikinase (EC 2.7.9.1). In segments taken from 4- and 8-day light-grown seedlings, BA did not enhance the photosynthetic activity nor the chlorophyll accumulation. The activity of the enzymes mentioned above, was significantly enhanced by the BA-treatment. BA mainly affected grana stacking in mesophyll cell chloroplasts in primary leaf segments taken from 3- to 5-day light-grown seedlings. Stroma thylakoid development was stimulated only in leaf segments from 3-day-old plants. At the same time BA accelerated grana loss in chloroplasts of bundle sheath cells, a typical phenomenon of development in such chloroplasts. Stroma thylakoid length in these chloroplasts increased by a BA treatment in segments from 3- and 4-day light-grown plants. A significantly higher number of chloroplasts was only observed with segments taken from 8-day light-grown seedlings and treated with BA. The etiochloroplast number in segments taken from 8-day etiolated plants was significantly higher in BA-treated segments after 26 h illumination. In etiochloroplasts from both mesophyll and bundle sheath cells, BA enhanced grana stacking after illumination for 4 h or more, whereas stroma membrane length was significantly higher only after 26 h light. It is concluded that the effects of BA depend on the developmental stage. BA accelerates the development of mesophyll and bundle sheath cell (etio)chloroplasts, but does not affect the ultrastructure of mature chloroplasts.     

The occurrence of both C3 and C4 photosynthetic characteristics in a single Zea mays plant

The activities of the carboxylating enzymes ribulose-1,5-biphosphate (RuBP) carboxylase and phosphoenolpyruvate (PEP) carboxylase in leaves of three-week old Zea mays plants grown under phytotron conditions were found to vary according to leaf position. In the lower leaves the activity of PEP carboxylase was lower than that of RuBP carboxylase, while the upper leaves exhibited high levels of PEP carboxylase. Carbon dioxide compensation points and net photosynthetic rates also differed in the lower and upper leaves. Differences in the fine structure of the lowermost and uppermost leaves are shown. The existence of both the C₃ and C₄ photosynthetic pathways in the same plant, in this and other species, is discussed.Abbreviations PEP phosphoenolpyruvate - RuBP ribulose-1,5-biphosphate     

Extreme cytokinesis phenomenon in the phenotype of meiotic mutation pam of Zea mays

Our study of cytological phenotype of meiotic mutation pam resulted in detecting a failure of cytokinesis in mutant pollen mother cells in the form of a block of fusion of membrane vesicles of the cell plate, and an impossibility of formation of daughter cell membranes. The mutation does not disturb the division spindle structure and function. Asynchrony of meiosis in pam is the result of arrest of pollen mother cells at metaphase 1 and metaphase 2.     

A possible role for C4 photosynthetic enzymes in tolerance of Zea mays to NaCl

Treatment of 14-day-old maize cultivars (Hybrid351 and Giza2) with 250 mM NaCl significantly reduced shoot fresh and dry weights and protein content during the subsequent 12 days. The magnitude of reduction was more pronounced in Giza than Hybrid. Both cultivars contained converging levels of protein for the enzymes phosphoenolpyruvate carboxylase (PEPC), malate dehydrogenase (MDH), pyruvate phosphate dikinase (PPDK) and ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) under normal conditions; however, NaCl led to increase these levels in Hybrid and decrease them in Giza. Moreover, NaCl significantly inhibited the activities of PEPC, MDH and PPDK in both cultivars during the first 2 days, thereafter the inhibition nullified only in Hybrid; nonetheless, Rubisco was the least affected enzyme in both cultivars. In addition, NaCl slightly increased V _max of PEPC, MDH and PPDK in Hybrid with no change in K _m; nevertheless V _max dropped in Giza with an increase in K _m of only PEPC and MDH. Also K _cat, K _cat/K _m and V _max/K _m of all enzymes were lower in treated Giza than in treated Hybrid. The increased V _max of all enzymes in only Hybrid by NaCl confirms that they were synthesised more in Hybrid than in Giza. However, the decreased V _max in Giza concomitant with the increased K _m points to an interference of salinity with synthesis of enzymes and their structural integrity. This would lead to a noncompetitive inhibition for the enzymes. These findings declare that maize tolerance to NaCl was larger in Hybrid compared to Giza due to a role for C4 enzymes.     

The Hydrolysis of Endosperm Protein in Zea mays

Degradation of the major storage proteins in maize endosperm, zein and glutelin, begins during the 2nd day of germination. The protein most abundant in the mature endosperm is degraded most rapidly. The patterns of protein loss are essentially similar in germinating seeds and excised endosperms. Cycloheximide, added at the beginning of the incubation period, prevents the development of α-amylase and protease activities and the disappearance of starch and protein reserves. Late additions (70 hours) of cycloheximide still inhibit the increase in hydrolase activity but have no effect on the hydrolysis of storage reserves. The results indicate that the hydrolytic enzymes are synthesized de novo in the maize endosperm.     

弱光胁迫影响夏玉米光合效率的生理机制初探

大田条件下, 以普通夏玉米(Zea mays) ‘泰玉2号’为材料, 于授粉后1-20天遮光55% (+S), 以大田自然光照条件下生长的玉米作为对照(-S), 研究了遮光及恢复过程中玉米植株的光合性能、叶绿体荧光参数、叶黄素循环以及光能分配的变化, 初步揭示夏玉米开花后弱光条件下光适应的生理机制, 为玉米高产稳产提供理论依据。结果表明, 遮光后玉米穗位叶叶绿素含量及可溶性蛋白含量均减少, RuBP羧化酶和PEP羧化酶活性显著降低, 导致穗位叶净光合速率(P_n)迅速下降, 光饱和点也明显降低; 恢复初期P_n迅速升高, 光合关键酶活性有所增强。遮光后植株的最大光化学效率(F_v/F_m)、实际光化学效率(Ф_PSII)显著降低, 非光化学淬灭(NPQ)则显著升高, 而恢复初期植株穗位叶Ф_PSII有所升高, 表明突然暴露在自然光下的光合电子传递速率明显加快, 这与其光合速率及光合酶活性的趋势保持一致; 遮光处理对穗位叶叶黄素循环库的大小(紫黄质+花药黄质+玉米黄质(V + A + Z))影响不显著, 但使叶黄素循环的脱环氧化状态(A + Z)/(V + A + Z)增加; 遮光后植株分配于光化学反应的光能明显减少, 天线耗散光能比率显著增加, 恢复过程中植株主要以过剩非光化学反应的形式耗散过剩的光能。遮光后及恢复初期, 玉米植株的PSII原初光化学活性明显下降, 限制了光合碳代谢的电子供应从而抑制了光合作用, 主要依赖叶黄素循环途径进行能量耗散, 而在光照转换后遮光的玉米叶片在适应自然光过程中的光保护机制不断完善, 光合能力逐渐得到 恢复。     

Site-specific recombination in Zea mays

The elimination of marker genes after selection is recommended for the commercial use of genetically modified plants. We compared the applicability of the two site-specific recombination systems Cre/lox and Flp/FRT for marker gene elimination in maize plants. The selection marker gene pat surrounded by two identically directed lox or FRT sites was introduced into maize. Sexual crossing with plants harboring the corresponding constitutively expressed recombinase led to the precise and complete excision of the lox-flanked marker gene in the F₁ progeny, whereas Flp-mediated recombination of FRT sequences occurred rarely. Further examination of site-specific integration was done by biolistic bombardment of immature embryos harboring only one lox site with a lox.uidA sequence with results indicating directed integration.