Developmental changes of plant affecting primary photosynthate distribution in rice leaves

Developmental changes of plant in the regulation of photosynthate distribution of leaves were studied in hydroponically cultivated rice by the ¹⁴CO₂ tracer technique and analysis of the activity of the regulatory enzymes, sucrose phosphate synthase (SPS), phosphoenolpyruvate carboxylase (PEPC), and pyruvate kinase (PK). The distribution of primary photosynthates into sugars, amino acids, organic acids, sugar phosphates, proteins, and polysaccharides was determined by column chromatography. The relative primary photosynthate distribution to the sugar phosphate fraction was significantly larger in the 5^th leaf than in the 6^th one. Correspondingly, the V_max of PEPC was significantly higher in the 5^th than in the 6^th leaf, while no significant differences between leaves were detected in the other enzymes. As a consequence, the ratio of the V_max of SPS and PEPC was lower in the 5^th than in the 6^th leaf. As the 5^th leaf develops before panicle initiation in rice, it predominantly supports vegetative growth, while the 6^th leaf develops after panicle initiation and thus contributes mainly to reproductive growth. We conclude that the physiological properties of each leaf are regulated developmentally. When the 6^th leaf became fully expanded (corresponding to the panicle initiation stage of plant), the distribution pattern of ¹⁴C was transiently changed in the 5^th leaf, indicating that individual organs that are mainly involved in vegetative development are affected to some extent by the whole-plant-level physiological transformation that occurs at the transition from the vegetative to the reproductive stage.     

Photosynthetic Enzymes of the C4 Grass Setaria sphacelata Under Water Stress: A Comparison Between Rapidly and Slowly Imposed Water Deficit

Two stress imposing systems were used: a rapid stress developed by allowing excised leaves to loose water by transpiration, and a slow stress developed by withholding watering of potted plants. Carboxylating enzymes reacted differently on both types of stress. Rapid stress increased ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) activation, but both activities (initial and total) showed little variation with stress. Under slow stress the activation did not change, although both activities decreased much under stress. Phosphoenolpyruvate carboxylase (PEPC) showed a deep decrease of activity under rapid stress, nevertheless, a certain recovery was found under extreme stress. On the other hand, under slow stress the activity of PEPC showed a linear increase with decreasing relative water content. The ratio between physiological and maximal activity increased slightly under both types of stress. The activity of malic enzyme did not change under rapid stress, and decreased linearly under slow stress.     

Effect of Chronic O3 Fumigation on the Activity of Some Calvin Cycle Enzymes in Two Poplar Clones

The effects of long-term exposure to ozone (O₃, 60 mm³ m^-3 for 5 h d^-1) on some Calvin cycle enzymes, in particular those modulated by the thioredoxin system, were studied in two poplar clones. These clones differ in sensitivity to O₃. In the I-214 clone, the first effects from O₃ treatment were seen after 40 d of fumigation, while the Eridano clone showed visible symptoms of damage after only 15 d of the treatment. Specific activity of ribulose-1,5-bisphosphate carboxylase/oxygenase (E.C. 4.1.1.39) diminished in both the clones, while specific activity of phosphoenolpyruvate carboxylase (E.C. 4.1.1.31) increased. Exposure to O₃ also caused a reduction in the specific activity of ribulose-1,5-bisphosphate kinase (E.C. 2.7.1.19) in both clones. At the end of the exposure to O₃, specific activity of glyceraldehyde 3-phosphate dehydrogenase (E.C. 1.2.1.13) increased in I-214 and remained similar to the control in Eridano, whereas specific activity of fructose-1,6-bisphosphate phosphatase (E.C. 3.1.3.11) was higher in Eridano and similar to the control in I-214.     

Changes of Photosynthetic Characteristics in Relation to Leaf Senescence in Two Maize Hybrids with Different Senescent Appearance

A field experiment was conducted to investigate the changes in chlorophyll (Chl) and nitrogen (N) contents, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) and phosphoenolpyruvate carboxylase (PEPC) contents and PEPC activity, and the photon-saturated net photosynthetic rate (P _Nsat), and their relationships with leaf senescence in two maize hybrids with different senescent appearance. One stay-green (cv. P3845) and one earlier senescent (cv. Hokkou 55) hybrid were used in this study, and we found that Chl and N contents and the P _Nsat in individual leaves of P3845 were greater than those in corresponding leaves of Hokkou 55 at the successive growth stages. In addition, larger contents of RuBPCO and PEPC, and a greater activity of PEPC were observed in P3845. Due to the lower rates of decrease of Chl, RuBPCO, and PEPC amounts per unit of N, and the lower net C translocation rate per unit of N in the stay-green hybrid, leaf senescence was delayed in comparison to the earlier senescent hybrid.     

Net CO2 uptake rates for Hylocereus undatus and Selenicereus megalanthus under field conditions: Drought influence and a novel method for analyzing temperature dependence

Net CO₂ uptake rates (P _N) were measured for the vine cacti Hylocereus undatus and Selenicereus megalanthus under relatively extreme climatic conditions in Israel. Withholding water decreased rates and the daily amount of CO₂ uptake by about 10 % per day. Compared with more moderate climates within environmental chambers, the higher temperatures and lower relative humidity in the field led to a more rapid response to drought. The upper envelopes of scatter diagrams for P _N versus temperature for these Crassulacean acid metabolism species, which indicate the maximal rates at a particular temperature, were determined for both night time CO₂ uptake in Phase I (mediated by phosphoenolpyruvate carboxylase, PEPC) and early morning uptake in Phase II (mediated by ribulose-1,5-bisphosphate carboxylase/oxygenase, RuBPCO). As stem temperature increased above 13 °C, the maximal P _N increased exponentially, reaching maxima near 27 °C of 12 and 8 μmol m⁻² s⁻¹ for Phases I and II, respectively, for H. undatus and 6 and 4 μmol m⁻² s⁻¹, respectively, for S. megalanthus. Based on the Arrhenius equation, the apparent activation energies of PEPC and RuBPCO were 103 and 86 kJ mol⁻¹, respectively, for H. undatus and 77 and 49 kJ mol⁻¹, respectively, for S. megalanthus, within the range determined for a diverse group of species using different methodologies. Above 28 °C, P _N decreased an average of 58 % per °C in Phase I and 30 % per °C in Phase II for the two species; such steep declines with temperature indicate that irrigation then may lead to only small enhancements in net CO₂ uptake ability.     

Altitude-related changes in activities of carbon metabolism enzymes in <Emphasis Type="Italic">Rumex nepalensis</Emphasis>

Activities of some enzymes related to carbon metabolism were studied in different ecotypes of Rumex nepalensis growing at 1 300, 2 250, and 3 250 m above mean sea level. Activities of ribulose-1,5-bisphosphate carboxylase/oxygenase, phosphoenolpyruvate carboxylase, aspartate aminotransferase, and glutamine synthetase increased with altitude, whereas activities of malate dehydrogenase, NAD-malic enzyme, and citrate synthase did not show a significant difference with change in altitude.     

Abscisic acid promoted changes in the protein profiles of rice seedling by proteome analysis

This study investigates the influence of exogenously applied abscisic acid (ABA) on the leaves and leaf sheaths of two-week-old rice seedling at the level of the proteome. Significant differences were observed in the two-dimensional polyacrylamide gel electrophoresis protein profiles between control and ABA treated samples. Amino-acid sequence analysis of affected proteins revealed that ABA caused drastic changes in the major photosynthetic protein, ribulose 1,5-bisphosphate carboxylase/oxygenase and accumulation of certain defense/stress-related proteins. Moreover, cutting or treating leaf sheaths with jasmonic acid (JA) rapidly increased the endogenous level of ABA, suggesting a role for ABA during the defense/stress-response. Comparative study indicated a potential overlap between ABA and JA as detected at the level of the proteome. Furthermore, in vitro protein phosphorylation experiments and in-gel kinase assays also revealed considerable changes in the phosphorylation status of some proteins, and differential effects on myelin basic protein and calcium-dependent protein kinase activities by ABA treatment, which suggests involvement of kinase in the downstream signaling cascade. These results provide evidence at proteome level for the involvement of ABA in stress-response in rice seedling.     

Elevated CO<Subscript>2</Subscript> and temperature differentially affect photosynthesis and resource allocation in flag and penultimate leaves of wheat

Differences in acclimation to elevated growth CO₂ (700 μmol mol⁻¹, EC) and elevated temperature (ambient +4 °C, ET) in successive leaves of wheat were investigated in field chambers. At a common measurement CO₂, EC increased photosynthesis and the quantum yield of electron transport (Φ) early on in the growth of penultimate leaves, and later decreased them. In contrast, EC did not change photosynthesis, and increased Φ at later growth stages in the flag leaf. Contents of chlorophyll (Chl), ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO), and total soluble protein were initially higher and subsequently lower in penultimate than flag leaves. EC decreased RuBPCO protein content relative to soluble protein and Chl contents throughout the development of penultimate leaves. On the other hand, EC initially increased the RuBPCO:Chl and Chl a/b ratios, but later decreased them in flag leaves. In the flag leaves but not in the penultimate leaves, ET initially decreased initial and specific RuBPCO activities at ambient CO₂ (AC) and increased them at EC. Late in leaf growth, ET decreased Chl contents under AC in both kinds of leaves, and had no effect or a positive one under EC. Thus the differences between the two kinds of leaves were due to resource availability, and to EC-increased allocation of resources to photon harvesting in the penultimate leaves, but to increased allocation to carboxylation early on in growth, and to light harvesting subsequently, in the flag leaves.     

Extent of ipt gene expression and resulting amount of cytokinins affect activities of carboxylation enzymes in transgenic plants

Three types of transgenic plants of Solanum tuberosum cvs. Kamyk and Oreb, and Nicotiana tabacum cvs. Maryland Mammoth and Trapezond were selected according to intensity of introduced ipt gene expression and resulting amount of synthesised cytokinins (CKs). In comparison with controls, original transgenic regenerants grown in vitro showed a massive increase of CK contents, in tobacco by 379 % and in potato by 159 % (MAS). Potato grown in soil from tubers of transgenic plants demonstrated a moderate increase (44 %) of CK contents (MOD). Transgenic tobacco grown from seeds in vitro did not show any significant change in CK contents (NOT). Initial (RuBPC_i and RuBPO_i) and total (RuBPC_t) activities of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO), and the activity of phosphoenolpyruvate carboxylase (PEPC) were not significantly affected by the transformation in the NOT plants. In the MOD plants, the RuBPCO activities were stimulated by up to 34 % whereas the PEPC activity was decreased by 17 %. On the other hand, all the measured enzyme activities were 32 – 91 % lower in the MAS. Leaf area, fresh and dry masses, and chlorophyll and soluble protein contents also went down with increasing CK amounts in the transformants. Dependence of RuBPC_i/RuBPO_i and RuBPC_t/PEPC ratios on the relative CK amounts in transgenic plants revealed that the individual enzyme activities were not affected uniformly. Endogenous CK contents in the MAS thus apparently exceeded an optimum needed for positive effects on many physiological traits and became a stress factor for such plants.     

ABA modulates the degradation of phosphoenolpyruvate carboxylase kinase in sorghum leaves

Salt stresses strongly enhance the phosphoenolpyruvate carboxylase kinase (PEPC-k) activity of sorghum leaves. This work shows that (1) abscisic acid (ABA) increased the rise in kinase activity in illuminated leaf disks of the non-stressed plant, (2) ABA decreased the disappearance of PEPC-k activity in the dark, (3) two PEPC-k genes expressed in sorghum leaves, PPCK1 and PPCK2, were not up-regulated by the phytohormone and, (4) ABA effects were mimicked by MG132, a powerful inhibitor of the ubiquitin-proteasome pathway. Collectively these data support a role for the ubiquitin-proteasome pathway in the rapid turnover of PEPC-k. The negative control by ABA on this pathway might account for the increase of kinase activity observed in salt-treated plants.     

Photosynthetic characteristics of C4 trait in chlorina mutant of rice (Oryza sativa L.)

Biao 810S is a chlorina mutant of the thermosensitive genic male sterile (TGMS) rice. We compared photosynthetic characteristics of these two lines. The contents of chlorophylls and carotenoids in Biao 810S were approximately half of those in 810S. However, the net photosynthetic rate (P _N) of Biao 810S was higher than that of 810S under high irradiance or low concentration of carbon dioxide, and the photon quantum efficiency was higher than that of 810S. The activity of ribulose-1,5-bisphosphate carboxylase/oxygenase in Biao 810S was only 69.80 % of that in 810S, but the activities of phosphoenolpyruvate carboxylase and NADP-malic enzyme were 79.50 and 69.06 % higher than those of 810S, respectively, suggesting that the efficiency of photon energy utilization in Biao 810S was enhanced by reduction of thermal dissipation and increase of electron transfer rate to generate sufficient assimilation power for the dark reactions. Consequently, the increased activities of C₄ photosynthetic enzymes lead to more effective fixation of CO₂ and the synergistic effect of light and dark reactions contributed to the higher P _N of Biao 810S.     

Comparative proteomic study reveals dynamic proteome changes between superhybrid rice LYP9 and its parents at different developmental stages

Heterosis is a common phenomenon in which the hybrids exhibit superior agronomic performance than either inbred parental lines. Although hybrid rice is one of the most successful apotheoses in crops utilizing heterosis, the molecular mechanisms underlying rice heterosis remain elusive. To gain a better understanding of the molecular mechanisms of rice heterosis, comparative leaf proteomic analysis between a superhybrid rice LYP9 and its parental cultivars 9311 and PA64s at tillering, flowering and grain-filling stages were carried out. A total of 384 differentially expressed proteins (DP) were detected and 297 DP were identified, corresponding to 222 unique proteins. As DP were divided into those between the parents (DP_PP) and between the hybrid and its parents (DP_HP), the comparative results demonstrate that proteins in the categories of photosynthesis, glycolysis, and disease/defense were mainly enriched in DP. Moreover, the number of identified DP_HP involved in photosynthesis, glycolysis, and disease/defense increased at flowering and grain-filling stages as compared to that at the tillering stage. Most of the up-regulated DP_HP involved in the three categories showed greater expression in LYP9 at flowering and grain-filling stages than at the tillering stage. In addition, CO₂ assimilation rate and apparent quantum yield of photosynthesis also showed a greater increase in LYP9 at flowering and grain-filling stages than at the tillering stage. These results suggest that the proteins involved in photosynthesis, glycolysis, and disease/defense as well as their dynamic regulation at different developmental stages may be responsible for heterosis in rice.     

Changes of photosynthetic activities of maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) seedlings in response to cadmium stress

Maize (Zea mays L.) seedlings were grown in nutrient solution culture containing 0, 5, and 20 μM cadmium (Cd) and the effects on various aspects of photosynthesis were investigated after 24, 48, 96 and 168 h of Cd treatments. Photosynthetic rate (P _N) decreased after 48 h of 20 μM Cd and 96 h of 5μM Cd addition, respectively. Chl a and total Chl content in leaves declined under 48 h of Cd exposure. Chl b content decreased on extending the period of Cd exposure to 96 h. The maximum quantum efficiency and potential photosynthetic capacity of PSII, indicated by F_v/F_m and F_v/F_o, respectively, were depressed after 96 h onset of Cd exposure. After 48 h of 5μM Cd and 24 h of 20 μM Cd treatments, the activities of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.39) and phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) in the leaves started to decrease, respectively. We found that the limitation of photosynthetic capacity in Cd stressed maize leaves was associated with Cd toxicity on the light and the dark stages. However, Cd stress initially reduced the activities of Rubisco and PEPC and subsequently affected the PSII electron transfer, suggesting that the Calvin cycle reactions in maize plants are the primary target of the Cd toxic effect rather than PSII.     

Water Stress and Day-To-Day Variation in Apparent Photosynthetic Acclimation of Field-Grown Soybeans to Elevated Carbon Dioxide Concentration

Midday measurements of single leaf gas exchange rates of upper canopy leaves of soybeans grown in the field at 350 (AC) and 700 (EC) µmol(CO₂) mol^–1 in open topped chambers sometimes indicated up to 50 % higher net photosynthetic rates (P _N) measured at EC in plants grown at AC compared to EC. On other days mean P _N were nearly identical in the two growth [CO₂] treatments. There was no seasonal pattern to the variable photosynthetic responses of soybean to growth [CO₂]. Even on days with significantly lower P _N in the plants grown at EC, there was no reduction in ribulose-1,5-bisphosphate carboxylase/oxygenase, chlorophyll, or soluble protein contents per unit of leaf area. Over three years, gas exchange evidence of acclimation occurred on days when either soil was dry or the water vapor pressure deficit was high (n = 12 d) and did not occur on days after rain or on days with low water vapor pressure deficit (n = 9 d). On days when photosynthetic acclimation was evident, midday leaf water potentials were consistently 0.2 to 0.3 MPa lower for the plants grown at EC than at AC. This suggested that greater susceptibility to water stress in plants grown at EC cause the apparent photosynthetic acclimation. In other experiments, plants were grown in well-watered pots in field chambers and removed to the laboratory early in the morning for gas exchange measurements. In these experiments, the amount of photosynthetic acclimation evident in the gas exchange measurements increased with the maximum water vapor pressure deficit on the day prior to the measurements, indicating a lag in the recovery of photosynthesis from water stress. The apparent increase in susceptibility to water stress in soybean plants grown at EC is opposite to that observed in some other species, where photosynthetic acclimation was evident under wet but not dry conditions, and may be related to the observation that hydraulic conductance is reduced in soybeans when grown at EC. The day-to-day variation in photosynthetic acclimation observed here may account for some of the conflicting results in the literature concerning the existence of acclimation to EC in field-grown plants.     

Jasmonic acid-induced necrosis and drastic decreases in ribulose-1,5-bisphosphate carboxylase/oxygenase in rice seedlings under light involves reactive oxygen species

Leaves and leaf sheaths of rice (Oryza sativa L.) seedlings responded to 100 μmol/L jasmonic acid with necrotic lesions and browning, respectively. By SDS-PAGE and immunoblot analysis, light-, dose- and time-dependent drastic reductions in ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) were found in jasmonate-treated leaves and leaf sheaths. Cycloheximide, free radical scavengers, a metal chelator, and protease and photosystem II inhibitors significantly reversed decrease in RuBisCO protein. Polypeptides were induced in light-incubated jasmonate-treated leaf sheaths. Surprisingly in dark incubated leaves only, induction was blocked by cycloheximide, indicatingde novo synthesis. Photosystem II inhibitors, the metal chelators 1,10-phenanthroline and EDTA, and a protease inhibitor, tosyl-lysine chloromethyl ketone, also blocked polypaptide induction, whereas free radical scavengers were only partially effective. Results indicate that jasmonate action in rice may be mediated through inhibition or damage of photosynthesis via production of reactive oxygen species and leading to necrotic lesion formation.     

Photosynthetic response of P-deficient Gracilaria tenuistipitata under two different phosphate treatments

Phosphorus-deficient Gracilaria tenuistipitata Zhang et Xia was cultured for 15 days at two different inorganic phosphate (P_i) concentrations: 3 μM (low P_i treatmenl) or 30 μM phosphate (high P_i treatment). The amount of ribulose-l,5-bisphosphate carboxy-lase/oxygenase (Rubisco), phycobiliproteins, Chl a and total soluble proteins were higher in the high P_i than in the low P_i treatment. The total N content of the low P_i plants was lower than in plants grown at high P_i concentrations whereas the amount of total C was highest in low P_i plants. The increase of Rubisco content in the high P_i treatment (3-fold) was parallel to the enhancement of the maximum photosynthetic rate which increased 5-fold. This correspondence was also found in the low P_i treatment in which Rubisco content and maximum photosynthesis did not change significantly from the initial values. The photosynthetic efficiency was also higher at high than at low P_i. The high P_i plants also showed higher dark respiration and growth rates. The data presented here suggest that marine macroalgae submitted to P_i deficiency exhibit a decrease in growth caused not only by P_i implication on energy transfer in photosynthesis and respiration, but also by the diminution of the amount of photosynthetic pigments and Rubisco.     

Regulation of Rubisco by inhibitors in the light

2-carboxy-D-arabinitol-1-phosphate (CA1P) bound to Rubisco either in leaf extracts or after purification can be displaced by SO₄^2? ions. Thus, treatment of leaf extracts with a buffer containing 200 mol m^?3 SO₄^2? displaces any bound CA1P and enables measurement of maximum car-boxylation potential. In tobacco leaves, the activity following treatment with SO₄^?2 ions (‘maximal activity’) is greater than the total Rubisco activity. The ratio of the two activities altered in a dynamic way with fluctuations in irradiance. Even in species which do not produce significant amounts of CA1P, the maximal activity greatly exceeded the total activity. Anion exchange separation of components in acid extracts confirmed the absence of CA1P in tobacco leaves harvested above an irradiance of 300 μmol quanta m^?2 s^?1, but the presence of another inhibitor of Rubisco. These results are consistent with the regulation of Rubisco activity by inhibitors other than CA1P which, like CA1P, can be displaced by SO₄^2? ions.     

Activity regulation and physiological impacts of maize C4-specific phosphoenolpyruvate carboxylase overproduced in transgenic rice plants

Phosphoenolpyruvate carboxylase (PEPC) was overproduced in the leaves of rice plants by introducing the intact maize C₄-specific PEPC gene. Maize PEPC in transgenic rice leaves underwent activity regulation through protein phosphorylation in a manner similar to endogenous rice PEPC but contrary to that occurring in maize leaves, being downregulated in the light and upregulated in the dark. Compared with untransformed rice, the level of the substrate for PEPC (phosphoenolpyruvate) was slightly lower and the product (oxaloacetate) was slightly higher in transgenic rice, suggesting that maize PEPC was functioning even though it remained dephosphorylated and less active in the light. ¹⁴CO₂ labeling experiments indicated that maize PEPC did not contribute significantly to the photosynthetic CO₂ fixation of transgenic rice plants. Rather, it slightly lowered the CO₂ assimilation rate. This effect was ascribable to the stimulation of respiration in the light, which was more marked at lower O₂ concentrations. It was concluded that overproduction of PEPC does not directly affect photosynthesis significantly but it suppresses photosynthesis indirectly by stimulating respiration in the light. We also found that while the steady-state stomatal aperture remained unaffected over a wide range of humidity, the stomatal opening under non-steady-state conditions was destabilized in transgenic rice. This revised version was published online in August 2006 with corrections to the Cover Date.