PEPCase Activity Research Articles

Coronatine enhances drought tolerance via improving antioxidative capacity to maintaining higher photosynthetic performance in soybean

Coronatine (COR), a structural and functional mimic of jasmonates, is involved in a wide array of effects on plant development and defence responses. This study was conducted to explore the role of exogenously applied COR in alleviating the adversities of drought stress in soybean. COR treatment markedly increased the activities of antioxidant enzymes and proline content, and reduced the accumulation of malondialdehyde and hydrogen peroxide under drought stress. Thus, COR-treated plants had higher leaf relative water content and lower electrolye leakage, which led to higher chlorophyll content, activities of RuBPCase and PEPCase, and net photosynthetic rate compared to control plants exposed to drought. COR also increased maximal efficiency of PS II photochemical reaction and photochemical quenching coefficient, but decreased non-photochemical quenching coefficient. These beneficial effects led to enhanced photosynthetic performance and the translocation of assimilated 14C which promoted growth and accumulation of dry biomass in COR-treated soybean plants subjected to drought. Interestingly, COR application did not affect the growth and biomass accumulation under well-watered condition. These results suggested the involvement of COR on improving drought tolerance in soybean by modulating antioxidant systems and membrane stability to maintain higher photosynthetic performance.

Exogenous glycinebetaine and humic acid improve growth, nitrogen status, photosynthesis, and antioxidant defense system and confer tolerance to nitrogen stress in maize seedlings

Abiotic stresses, including nitrogen stress (NS), can hamper photosynthesis and cause oxidative damage to plants. Upregulation of the antioxidative defense system and photosynthesis induced by exogenous glycinebetaine (GB) and humic acid (HA) can mitigate the inhibitory effects of NS on plants. In the present investigation, the beneficial effects of exogenously applied GB and HA were examined on growth, leaf N status, photosynthesis, lipid peroxidation, and activities of some key antioxidant enzymes in the seedlings of maize cv. Zhengdan 958 (ZD958) exposed to NS. NS caused a significant reduction in total dry matter of seedlings of ZD958, but both GB and HA proved effective in mitigating this inhibition, hence, the beneficial effects of GB being more pronounced than those of HA. NS led to a considerable decrease in leaf total N and endogenous GB contents, stomatal conductance (gs), net photosynthetic rate (Pn), intercellular CO2 concentration (Ci), and activities of two key C4 photosynthesis enzymes phosphoenolpyruvate carboxylase (PEPCase) and ribulose-1,5-bisphosphate carboxylase (RuBPCase) as well as of superoxide dismutase (SOD) and peroxidase (POD). This treatment caused an increase in lipid peroxidation, but showed no effect on POD activity. Exogenous application of varying doses of GB resulted in a decrease in lipid peroxidation and Ci, and an increase in leaf total N and endogenous glycinebetaine (EGB) content, Pn, and activities of RuBPCase, PEPCase, SOD, and catalase (CAT) under NS. In contrast, application of different doses of HA resulted in a decrease in lipid peroxidation, an increase in Pn, gs, and Ci as well as SOD, CAT, and POD activities without increasing leaf total N and EGB content, and enhanced RuBPCase and PEPCase activities. The present study suggests that exogenous application of GB and HA can induce tolerance in maize plants to NS, but through the regulation of different mechanisms.

Bicarbonate blocks iron translocation from cotyledons inducing iron stress responses in Citrus roots

The effect of bicarbonate ion (HCO3−) on the mobilization of iron (Fe) reserves from cotyledons to roots during early growth of citrus seedlings and its influence on the components of the iron acquisition system were studied. Monoembryonic seeds of Citrus limon (L.) were germinated “in vitro” on two iron-deprived media, supplemented or not with 10mM HCO3− (−Fe+Bic and −Fe, respectively). After 21d of culture, Fe concentration in seedling organs was measured, as well as gene expression and enzymatic activities. Finally, the effect of Fe resupply on the above responses was tested in the presence and absence of HCO3− (+Fe+Bic or +Fe, respectively). −Fe+Bic seedlings exhibited lower Fe concentration in shoots and roots than −Fe ones but higher in cotyledons, associated to a significative inhibition of NRAMP3 expression. HCO3− upregulated Strategy I related genes (FRO1, FRO2, HA1 and IRT1) and FC-R and H+-ATPase activities in roots of Fe-starved seedlings. PEPC1 expression and PEPCase activity were also increased. When −Fe+Bic pre-treated seedlings were transferred to Fe-containing media for 15d, Fe content in shoots and roots increased, although to a lower extent in the +Fe+Bic medium. Consequently, the above-described root responses became markedly repressed, however, this effect was less pronounced in +Fe+Bic seedlings. In conclusion, it appears that HCO3− prevents Fe translocation from cotyledons to shoot and root, therefore reducing their Fe levels. This triggers Fe-stress responses in the root, enhancing the expression of genes related with Fe uptake and the corresponding enzymatic activities.

Relationships Between C4 Enzyme Activities and Yield in Soybeans (Glycine max (L.) Merr.)

To study the relationships between C4 enzyme activities and yield, C4 enzyme activities (phosphoenolpyruvate carboxylase (PEPCase), NADP-malate dehydrogenase (NADP-MDH), NADP-malic enzyme (NADP-ME), and pyruvate phosphate dikinase (PPDK)) in different organs of ten soybean cultivars with different yields were measured at different growth stages in China. The result showed that four enzyme activities in C4 pathway were obviously different among cultivars, especially PPDK activity was not detected in the leaves of Dongnong 1567 and Dongnong 1068 and the young leaves of Gongjiao 9107-1 and Dongnong 97–172, but there were weak activities in pod coats. The order of C4 enzyme activities is young leaves < old leaves < pod coats. The correlation coefficients between PEPCase activity and yield and between NADP-MDH activity at blooming stage and yield were 0.6979 and 0.6565, respectively, and both reached the significant level (5%), and PEPCase activity kept significant positive correlation with plant photosynthetic rate. There was a negative correlation between NADP-ME activity and yield, and no correlation was found between PPDK activity and yield.

Expression of Maize Gene Encoding C4-Pyruvate Orthophosphate Dikinase (PPDK) and C4-Phosphoenolpyruvate Carboxylase (PEPC) in Transgenic Arabidopsis

To improve the efficiency of CO2 fixation in C3 photosynthesis, C4-cycle genes were overexpressed in Arabidopsis plants. In this study, a full-length cDNA for pyruvate orthophosphate dikinase (PPDK) with 971 amino acids was isolated from Zea mays using LA-PCR. The PPDK gene (GenBank accession number GU363532) and the phosphoenolpyruvate carboxylase (PEPC) gene (GenBank accession number: FJ415327) derived from a previous study were introduced into wild-type Arabidopsis both individually and in combination. The vector pCAMBIA3301 (p3301) was introduced into Arabidopsis as a control using an Agrobacterium-mediated system. The integration, transcription, and translation of the PPDK and the PEPC genes were confirmed by Southern blot, quantitative real-time PCR, and Western blot analysis. The average enzyme activity of C4-PPDK and C4-PEPC peaked at 4.63- and 5.81-fold compared to that of controls, respectively. The insertion of PPDK was found to increase the activity of PEPCase and vice versa. The photosynthetic rates of C4-PPDK, C4-PEPC, and C4-PKC(PPDK+PEPC) transgenic lines increased to 123.3, 136.4, and 146.6 % compared to that of controls, respectively.

Low dose of gamma irradiation enhanced drought tolerance in soybean

Drought stress is the main limiting factor in soybean production. However, no work has been done on how the application of a low dose of gamma rays could help to overcome water deficits during critical stages of soybean development. Gamma rays at a dose of 20 Gray (Gy) were applied to dry seeds of soybean before planting. Two levels of soil moisture (80% field capacity for well-watered control and 35% for drought-stressed treatment) were applied at pod initiation. Gamma irradiation increased biomass accumulation and seed yield in both treatments. It also increased the chlorophyll content, photosynthetic activity (14CO2fixation) and leaf water potential and enhanced the enzyme activities of RuBPCase and PEPCase of control plants compared with drought-stressed plants. Gamma irradiation (20 Gy) increased the soluble sugars, protein and proline content and the activities of peroxidase and superoxide dismutase in drought-stressed soybean leaves. It also increased the chloroplast size, which was reduced by drought treatment, and rebuilt, to some extent, the chloroplast ultrastructure. However, it decreased the malondialdehyde concentration and the electrical conductivity of the leaves under drought stress. Overall, the results indicated that pre-treatment with gamma rays (20 Gy) to dry seeds of soybean before planting could be used to enhance drought tolerance and minimize the yield loss caused by water deficit.

Breeding of Marker-free Indica Three-lines Restorer with ZmC4Ppc by Marker-assisted Selection

Enhancing photosynthetic productivity of rice is always the key goal of researchers by to use exogenous C4 genes. The full-length of intact maize gene of C4-specific PEPCase (ZmC4Ppc) is 6781 bp that makes it difficult for marker-assisted selection (MAS) in rice. The specific marker of intact ZmC4Ppc was designed and termed as MRpc. MRpc was verified that it was a gene marker because ZmC4Ppc could be overexpressed at every developmental stage and it was used for MAS of indica restorer Shuhui881 with ZmC4Ppc and FPM881was gotten, which was tested for genetic background, PEPCase activity, net photosynthetic rate (Pn), general combining ability (GCA) and specific combining ability (SCA). The results indicated that FPM881, which similarity of genetic background had arrived at above 95%, showed a higher PEPCase activity and net photosynthetic rate than those of control. Analysis of yield components showed that some of the progenies carrying ZmC4Ppc had better GCA and SCA than those of the control. Then, six lines of FPM881 were planted in larger population by selfing and gotten selectable-marker free plants with target gene using MRpc and hpt primers. Our results indicated that introduction of ZmC4Ppc via MAS and its stable expression could increase grain yield of hybrid rice.

Leaf anatomy and photosynthetic carbon metabolic characteristics in Phragmites communis in different soil water availability

To investigate the variations of anatomical and photosynthetic carbon metabolic characteristics within one species in response to increasing soil water stress, leaf anatomical characteristics, gas exchange and the activity of key enzymes in photosynthesis and photorespiration were compared in different ecotypes of Phragmitescommunis growing in an oasis-desert transitional zone (ODTZ) from swamp habitat (plot 1–3) via heavy salt meadow (plot 4–7) and light salt meadow habitat (plot 8–9) to dune habitat (plot 10–13) in Northwest China. The results showed that interveinal distance (ID) decreased with increasing water stress except that in plots of dune reed (DR). Vein mean diameter (VMD) in plot 10, 11 and 12 of the DR was significantly larger than that in other ecotypes. Leaf specific porosity (LSP) enhanced from plot 4 to plot 13 from heave salt meadow reed (HSMR) to light salt meadow reed (LSMR) and to DR. Chlorophyll fluorescence in bundle sheath cells were microscopically found in four ecotypes, especially significantly in the DR. Net CO2 assimilation rate (An) dropped rapidly from the swamp reed (SR) to the HSMR and then increased progressively from the LSMR to the DR. Stomatal conductance (gs) decreased and the water use efficiency (WUE) rose from the wet to the dry ecotypes. Sensitivity of gs to intercellular CO2 concentration (Ci) increased, but glycolate oxidase (GO) activity gradually reduced with increasing soil water deficiency. The RuBPCase activity did not reduce in four ecotypes even in DR, but, the PEPCase and NAD-ME activities as well as the ratio of PEPCase/RuBPCase were gradually enhanced with increasing soil water stress. We concluded that anatomical and photosynthetic carbon assimilating characteristics in P. communis were developing to the direction of C4 metabolism in response to the increasing drought stress in desert areas. The DR enduring severe water stress had more C4 like photosynthetic features than the HSMR and LSMR as well as SR, according to significantly increased VMD and LSP and higher gs sensitivity to Ci as well as higher PEPCase activity and lower GO activity in the DR.

The structural and photosynthetic characteristics of the exposed peduncle of wheat (Triticum aestivum L.): an important photosynthate source for grain-filling.

BackgroundIn wheat (Triticum aestivum L), the flag leaf has been thought of as the main source of assimilates for grain growth, whereas the peduncle has commonly been thought of as a transporting organ. The photosynthetic characteristics of the exposed peduncle have therefore been neglected. In this study, we investigated the anatomical traits of the exposed peduncle during wheat grain ontogenesis, and we compared the exposed peduncle to the flag leaf with respect to chloroplast ultrastructure, photosystem II (PSII) quantum yield, and phosphoenolpyruvate carboxylase (PEPCase; EC 4.1.1.31) activity.ResultsTransmission electron microscope observations showed well-developed chloroplasts with numerous granum stacks at grain-filling stages 1, 2 and 3 in both the flag leaf and the exposed peduncle. In the exposed peduncle, the membranes constituting the thylakoids were very distinct and plentiful, but in the flag leaf, there was a sharp breakdown at stage 4 and complete disintegration of the thylakoid membranes at stage 5. PSII quantum yield assays revealed that the photosynthetic efficiency remained constant at stages 1, 2 and 3 and then declined in both organs. However, the decline occurred more dramatically in the flag leaf than in the exposed peduncle. An enzyme assay showed that at stages 1 and 2 the PEPCase activity was lower in the exposed peduncle than in the flag leaf; but at stages 3, 4 and 5 the value was higher in the exposed peduncle, with a particularly significant difference observed at stage 5. Subjecting the exposed part of the peduncle to darkness following anthesis reduced the rate of grain growth.ConclusionOur results suggest that the exposed peduncle is a photosynthetically active organ that produces photosynthates and thereby makes a crucial contribution to grain growth, particularly during the late stages of grain-filling.

Nitrate uptake and utilization is modulated by exogenous γ-aminobutyric acid in Arabidopsis thaliana seedlings

Exogenously applied GABA modulates root growth by inhibition of root elongation when seedlings were grown in vitro on full-strength Murashige and Skoog (MS) salts, but root elongation was stimulated when seedlings were grown on 1/8 strength MS salts. When the concentration of single ions in MS salts was individually varied, the control of growth between inhibition and stimulation was found to be related to the level of nitrate (NO(3)(-)) in the growth medium. At NO(3)(-) concentrations below 40 mM (full-strength MS salts level), root growth was stimulated by the addition of GABA to the growth medium; whereas at concentrations above 40 mM NO(3)(-), the addition of GABA to the growth medium inhibited root elongation. GABA promoted NO(3)(-) uptake at low NO(3)(-), while GABA inhibited NO(3)(-) uptake at high NO(3)(-). Activities of several enzymes involved in nitrogen and carbon metabolism including nitrate reductase (NR), glutamine synthetase (GS), glutamate synthase (NADH-GOGAT), NADP-dependent isocitrate dehydrogenase (NADP-ICDH), and phosphoenol pyruvate carboxylase (PEPCase) were regulated by GABA in the growth medium. Supplementing 1/8 strength MS medium with 50 mM GABA enhanced the activities of all of the above enzymes except ICDH activities in root tissues. However, at full-strength MS, GABA showed no inhibitory effect on the activities of these enzymes, except on GS in both root and shoot tissues, and PEPCase activity in shoot tissues. Exogenous GABA increased the amount of NR protein rather than its activation status in the tissues. This study shows that GABA affects the growth of Arabidopsis, possibly by acting as a signaling molecule, modulating the activity of enzymes involved in primary nitrogen metabolism and nitrate uptake.

Response of cell surface pH to pCO 2 and iron limitation in the marine diatom Thalassiosira weissflogii

The physicochemical conditions immediately adjacent to the phytoplankton cell membrane, within the diffusion boundary layer, determine the speciation and reaction rates of biologically important nutrients such as iron and carbon dioxide. Over the past decade, several modeling efforts have attempted to calculate the supply of nutrients to phytoplankton surfaces using diffusion-reaction models. In these modeling efforts the pH within the cellular boundary layer is an important unknown and a direct measure of the cell surface pH is needed. We prepared and calibrated a new cell-bound, pH sensitive dye (a lectin–SNARF conjugate) that can be used to determine surface pH in living diatoms and applied the method to cultures grown under iron and carbon stress; conditions that are likely to elicit a pH response. We found that Thalassiosira weissflogii responded to iron limitation by reducing growth rates and increasing both the activity of phosphoenolpyruvate carboxylase (PEPCase) and the efflux of the organic acid malate, consistent with the iron-limited response of vascular plants termed Strategy I. Despite our predictions there is no proton efflux from iron or carbon stressed T. weissflogii and that the cell surface pH can be higher than the bulk seawater pH by as much as 0.4 units. High surface pH and increased PEPCase activity in low- pCO 2, high-iron grown cells is consistent with an efficient C4 type CO 2 concentrating mechanism. An increase in surface pH may impact cell surface precipitated iron and associated trace metals such as barium and may be an important factor to consider in paleo-export flux proxies when carbon dioxide is depleted.

Ozone effects on carbon metabolism in sensitive and insensitive Phaseolus cultivars

An ozone treatment of 165 nL L −1 for 3 h evoked differential responses in the primary leaves of two bean cultivars of the common bean ( Phaseolus vulgaris L.). While cv ‘Cannellino’ showed visible symptoms of injury within 24 h of exposure, no visible symptoms at all were evident in the cv ‘Top Crop’. In primary leaves of the sensitive cultivar Cannellino, we observed an increase in carbon breakdown (an increase in PFK and Fumarase) and a reduction in CO 2 photoassimilation, linked also to the diminished synthesis of sucrose (a decrease in SPS activity) and to the stimulation of the degradation of this sugar (an increase in SuSy and Invertase activities). A strong stimulation of PEPcase activity indicates both an increased synthesis of OAA and an enhanced replenishment of the tricarboxylic acid cycle. Finally, in Cannellino leaves the activity of NADP-malic enzyme increased indicating a stimulation of enzymes delivering NADPH. The findings of this research suggest that the visible symptoms in Cannellino represent an active response that this cultivar initiates to cope with excess oxidative load. The pattern in Top Crop was different. This cultivar did not show visible symptoms of injury, nor any responsive changes at the physiological and biochemical levels. Oxidative pathways are partially enhanced, e.g., increases in Invertase, PFK and IDH. There were also increases in some enzyme linked to the production of cytosolic NADPH as G6PD, probably caused by the slight increase in activity of the enzymes SKDH and PAL involved in synthesis of phenolic compounds. However, the absence of visible injury in Top Crop leaves is confirmation that, in this cultivar, the need to produce carbon skeleton and NADPH used in detoxification and repair process is lower.

Effects of Nitrate and Potassium on Ammonium Toxicity in Cucumber Plants

ABSTRACT Interactions between nitrate (NO3 −), potassium (K+), and ammonium (NH4 +) were investigated using hydroponically grown cucumber (Cucumis sativus L.) plants. Ammonium as the sole nitrogen (N) source at 10 mM was toxic and led to overall growth suppression, chlorosis, and necrosis of leaves. After 20 days, 50% of the plants were dead. However, when NO3 − was supplied at very low concentration together with high NH4 + (only 1% of total 10 mM N) all seedlings survived and their growth was improved. High K+ concentration (5 mM) also alleviated NH4 + toxicity and increased plant growth several fold compared to intermediate concentration of K+ (0.6 mM). Leaf total N and 15N derived from 15N-labelled NH4 + increased in the presence of NO3 −, but decreased at high K+ concentration. High K+ supply enhanced total carbon (C) and δ 13C and stimulated GS and PEPCase activities in leaves and roots. Nitrate supplementation had no effect on GS or PEPCase activities. It is concluded that K+ may alleviate NH4 + toxicity, partly by inhibiting NH4 + uptake, partly by stimulating C and N assimilation in the roots.

Effect of Ar+ Implantation and Maize Genome DNA on Autotetraploid Rice

The effect of Ar+ beam implantation and maize genome DNA on autotetraploid rice is studied. Better mutation types and higher mutation rates were discovered in M2 of T3 with ion implantation and immersion in maize genome DNA. In the five agronomic categories investigated, the mutation rate of the seed setting rate was 9.1%, and the total mutation rate was 14.8% in the T3. However, the total mutation rate was 2.1% with the treatment of only ion implantation and 1.3% with the treatment of only immersion in maize genome DNA. Mutant FA36(4) was discovered in M1 with ion beam implantation and immersion in maize genome DNA. Its RuBPCase activity, PEPCase activity and seed setting rate were 32%, 153%, and 36.79%, respectively, higher than its parent IR36(4). Rapid analysis of polymorphicDNA (RAPD) analysis of three M2 plants of FA36(4) (FM1, FM2, FM3) and two controls (purple maize and IR36(4)) were also conducted with 40 random primers. S5-3 was RAPD fragment amplified with a template of purple maize, FM2 and FM3 genome DNA using primer S5. There was no S5-3 in the RAPD pattern of IR36(4) or FM1.

Root Carbon Enrichment Alleviates Ammonium Toxicity in Cucumber Plants

ABSTRACT The addition of carbonates to a nutrient solution to alleviate ammonium (NH4 +) toxicity in hydroponically-grown cucumber (Cucumis sativus L.) plants was investigated. Stable isotopes [nitrogen (15N) and carbon (13C)] were used to assess the uptake of nitrogen [NH4 + or nitrate (NO3 −)] as well as carbon [bicarbonate (HCO3 −)/carbonate (CO3 2−)] by the roots. Ammonium as the sole N source at 5 mM decreased plant fresh weights compared to NO3 −. However, at lower concentrations of NH4 + (25% of 5 mM total N), growth was increased compared to NO3 − alone. Inorganic C enrichment [calcium carbonate (CaCO3)] of the nutrient solution increased the fresh weight of NH4 + grown plants with up to 150% relative to control plants receiving calcium hydroxide [Ca(OH)2] for pH regulation. Root 15N enrichment was lower in 15NH4 + supplied plants compared to 15NO3 −, while the 13C enrichment in leaves was increased by NH4 + nutrition compared to NO3 − or NH4NO3. The enhanced C capture was associated with high PEPCase activity in the roots. It is concluded that inorganic carbon enrichment of the root medium may alleviate NH4 + toxicity via increased synthesis of C skeletons and, accordingly, increased capacity for NH4 + assimilation and N export to the shoots.

Marker-assisted Selection of ZmC 4Ppc in Rice Breeding and Yield Trait Performances of Advanced Lines

The full-length of intact Zea mays gene for phosphoenolpyruvate carboxylase gene ( ZmC 4Ppc ) is 6 781 bp. The products of PCR for this gene were not clear with poor repeatability, resulting in that it was difficult for marker-assisted selection (MAS) both in rice and maize. For selecting the markers for MAS, sequences presented only in maize rather than in rice were identified by BLAST, and used for primer design using Primer Premier 5.0. A pair of specific primer termed MRpc (Forward: 5′ AAGCAGGGAAGCGAGACG 3′, Reverse: 5′ GATTGCCGCCAGCAGTAG 3′) was used for selection of transformed rice, and ZmC 4Ppc could be highly and constitutively expressed at each tested developmental stages in the transformed rice selected by using MRpc. Thus, MRpc was used for MAS of progenies carrying ZmC 4Ppc gene in rice and some restorer lines with ZmC 4Ppc (e.g. FPM881) derived from ZmC 4Ppc -transformed Kitaake backcrossed with a restorer line Shuhui 881 were obtained. The analyses on genetic background, PEPCase activity, net photosynthetic rate, general combining ability (GCA) and specific combining ability (SCA) of FPM881 showed that similarity of genetic background reached above 95%, the PEPCase and net photosynthetic rate were higher than those of the control, and some of the progenies carrying ZmC 4Ppc gene had better GCA and SCA for grain yield per plant, number of panicles per plant, and 1000-grain weight than those of the control. This suggested that the introduction of maize ZmC 4Ppc gene via MAS and its stable expression could increase grain yield of rice and would likely provide a pathway for rice varietal improvement.

Characteristics of ribulose-1,5-bisphosphate carboxylase and C4 pathway key enzymes in flag leaves of a super-high-yield hybrid rice and its parents during the reproductive stage

A newly-developed super-high-yield two-line hybrid rice ( Oryza sativa L.) LiangYouPeiJiu (LYPJ) together with its paternal line Wumang 9311 (WM9311) and maternal line Peiai 64S (PA64S) were used as experimental materials. The activities of C4 pathway photosynthetic enzymes, i.e. phosphoenolpyruvatecarboxylase (PEPcase), NADP-malic enzyme (NADP-ME), NADP-malate dehydrogenase (NADP-MDH), pyruvate phosphate dikinase (PPDK) and the key C3 pathway enzyme–ribulose-1,5-bisphosphate carboxylase (RuBPcase), as well as soluble protein were assayed during the days after emergence (DAE) of flag leaves. The results indicated that C4 pathway enzymes were expressed differently during the flag leaf's span of life. The activity of PEPcase, NADP-MDH, and PPDK were increased with leaf age after full expansion, reached the peak at around 32 DAE, and then fell down significantly, and the activity of NADP-ME increased before 23 DAE and decreased earlier than other C4 enzymes, at 32 DAE, indicating that NADP-ME may be more sensitive to senescence than other C4 enzymes. The activity of RuBPcase was significantly higher in LYPJ than that in its parents during the whole life span of flag leaves. The decrease in RuBPcase activity was greater than that in soluble protein content. The onset of loss of RuBPcase activity (11 DAE) occurred earlier than that of soluble protein content (23 DAE). In comparison, there was no significant difference in C4 pathway enzymes between LYPJ and its parents.

Awns play a dominant role in carbohydrate production during the grain‐filling stages in wheat (Triticum aestivum)

In wheat (Triticum aestivum L), the leaves particularly flag leaves have been considered to be the key organs contributing to higher yields, whereas awns have been considered subsidiary organs. Compared with extensive investigations on the assimilation contribution of leaves, the photosynthetic characteristics of awns have not been well studied. In this study, we investigated the ultrastructure of chloroplasts, oxygen evolution, and phosphoenolpyruvate carboxylase [phosphoenolpyruvate carboxylase (PEPCase) EC 4.1.1.31)] activity in both flag leaves and awns during the ontogenesis of wheat. Transmission electron microscope observations showed initial increases in the sizes of grana and the degree of granum stacks from the florescence‐emergence stage both in flag leaves and in awns, followed by the breakdown of membrane systems after the milk‐development stage. The results of oxygen evolution assays revealed that in both organs, the rate of photosynthesis increased in the first few stages and then decreased, but the decrease occurred much earlier in flag leaves than in awns. A PEPCase activity assay demonstrated that the activity of PEPCase was much higher in awns than in flag leaves throughout ontogeny; the value was particularly high at the late stages of grain filling. Our results suggest that awns play a dominant role in contributing to large grains and a high grain yield in awned wheat cultivars, particularly during the grain‐filling stages.

Effect of altitude on the primary products of photosynthesis and the associated enzymes in barley and wheat

There is little information available on the primary products of photosynthesis and the change in the activity of the associated enzymes with altitude. We studied the same in varieties of barley and wheat grown at 1300 (low altitude, LA) and 4200 m (high altitude, HA) elevations above mean sea level in the western Himalayas. Plants at both the locations had similar photosynthetic rates, leaf water potential and the chlorophyll fluorescence kinetics. The short-term radio-labelling experiments in leaves showed appearance of (14)CO(2) in phosphoglyceric acid and sugar phosphates in plants at both the LA and HA, suggesting a major role of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) in CO(2) fixation in the plants at two altitudes, whereas the appearance of labelled carbon in aspartate (Asp) and glutamate (Glu) at HA suggested a role of phosphoenolpyruvate carboxylase (PEPCase) in photosynthesis metabolism. Plants at HA had significantly higher activities of PEPCase, carboxylase and oxygenase activity of Rubisco, aspartate aminotransferase (AspAT), and glutamine synthetase (GS). However, the activities of malate dehydrogenase, NAD-malic enzyme and citrate synthase were similar at the two locations. Such an altered metabolism at HA suggested that PEPCase probably captured CO(2) directly from the atmosphere and/or that generated metabolically e.g. from photorespiration at HA. Higher oxygenase activity at HA suggests high photorespiratory activity. OAA thus produced could be additionally channelised for Asp synthesis using Glu as a source of ammonia. Higher GS activity ensures higher assimilation rate of NH(3) and the synthesis of Glu through GS-GOGAT (glutamine:2-oxoglutarate aminotransferase) pathway, also as supported by the appearance of radiolabel in Glu at HA. Enhanced PEPCase activity coupled with higher activities of AspAT and GS suggests a role in conserving C and N in the HA environment.

Temperature determines the occurrence of CAM or C3 photosynthesis in pineapple plantlets grown in vitro

Ananas comosus (L.) Merr. var. Smooth Cayenne plants when grown in vitro under different temperature regimes developed as CAM or as C3 plants. The plants used in this study were developed from the lateral buds of the nodal etiolated stem explants cultured on Murashige and Skoog medium for 3 mo. The cultures were maintained under a 16-h photoperiod for different thermoperiods. With 28°C light/15°C dark thermoperiod, as compared with constant 28°C light and dark, pineapple plants had a succulence index two times greater, and also a greater nocturnal titratable acidity and phosphoenolpyruvate carboxylase (PEPCase) activity, indicating CAM-type photosynthesis. The highest abscisic acid (ABA) level occurred during the light period, 8 h prior to maximum PEPCase activity, while the indole-3-acetic acid (IAA) peak was found during the dark period, coinciding with the time of highest PEPCase activity. These plants were also smaller with thicker leaves and fewer roots, but had greater dry weight. Their leaves showed histological characteristics of CAM plants, such as the presence of greater quantities of chlorenchyma and hypoderm. In addition, their vascular system was more conspicuous. In contrast, under constant temperature (28°C light/dark) plants showed little succulence in the leaves. There was no significant acid oscillation and diurnal variation in PEPCase activity in these plants, suggesting the occurrence of C3 photosynthesis. Also, no diurnal variation in ABA and IAA contents was observed. The results of this study clearly indicate a role for temperature in determining the type of carbon fixation pathway in in vitro grown pineapple. Evidence that ABA and IAA participate in CAM signaling is provided.