Effect of some heterocyclic herbicides on CO2 assimilation by spinach chloroplasts

We have studied the inhibitory effect of heterocyclic herbicides simazine, paraquat, pyrazon and amitrole on photosynthetic CO₂ fixation and on the level of intermediates of the CO₂ assimilation cycle in isolated spinach (Spinacia oleracea) chloroplasts, as well as their in vitro activities on ribulose-1,5-bisphosphate carboxylase and fructose-1,6-bisphosphatase. The half inhibitory concentrations (I₅₀) of CO₂ assimilation were about 1 μM for simazine and paraquat, and 10 μM for pyrazon. Amitrole, with an I₅₀ 100 μM, gave only a weak inhibition. In the presence of simazine or pyrazon the triose-phosphates/phosphoglycerate ratio diminished because of a decrease of the triose-phosphates percentage from 47% to 19%, which means an inhibition of the phosphoglycerate reduction step by a low NADPH synthesis. However, there was not a parallel increase of phosphoglycerate, because of collateral pathways leading to phospho-enolpyruvate, amino acids and other non-identified compounds. Paraquat did not give such a decreased ratio, which could be explained as an inhibition of some step of the Calvin cycle later than triose-phosphates by the H₂O₂ formed in a Mehler reaction. Amitrole did not show any effect on the pattern of intermediates. Simazine and pyrazon at 10 μM concentration promote a 20–30% activation of ribulose-1,5-bisphosphate carboxylase activity, whereas paraquat, pyrazon and simazine showed an I₅₀ about 100 μM for the inhibition of the photosynthetic fructose-1,6-bisphosphatase.     

Heterogeneity of leaf CO2 assimilation during photosynthetic induction

Spatial mid temporal variations in the distribution of photosynthesis over the leaf area were investigated during induction upon illumination of Rosa rubiginosa L. leaves. Gas exchange and maps of relative photosynthetie electron transport activity computed from chlorophyll fluorescence images were simultaneously monitored. In air, after 15 h of dark adaptation, linear electron transport was heterogeneously distributed over the leaf area during the induction. This patchy induction was explained by asynchronous metabolism activation for the first 10 min of illumination, concomitant asynchronous limitation by intrinsic metabolism and stomatal apertures (10–30 min) and finally by only stomatal limitation beyond 30 min. A brief transition to non-photorespiratory conditions after 20 min of illumination under subsaturating irradiance revealed a marked heterogeneity of CO₂ assimilation, presumably as a result of heterogeneous stomatal apertures. The frequency distribution of CO₂ assimilation was unimodal. During the induction, heterogeneity gradually decreased and photosynthesis was uniform at steady-state. After 10 min of dark adaptation, heterogeneity of linear electron transport activity occurred during the first 15 min of a second induction and mainly resulted from metabolic limitation.     

Decrease of phosphoribulokinase activity by antisense RNA in transgenic tobacco: definition of the light environment under which phosphoribulokinase is not in large excess

 To test the hypothesis that the contribution of phosphoribulokinase (PRK) to the control of photosynthesis changes depending on the light environment of the plant, the response of transgenic tobacco (Nicotiana tabacum L.) transformed with antisense PRK constructs to irradiance was determined. In plants grown under low irradiance (330 μmol m⁻² s⁻¹) steady-state photosynthesis was limited in plants with decreased PRK activity upon exposure to higher irradiance, with a control coefficient of PRK for CO₂ assimilation of 0.25 at and above 800 μmol m⁻² s⁻¹. The flux control coefficient of PRK for steady-state CO₂ assimilation was zero, however, at all irradiances in plant material grown at 800 μmol m⁻² s⁻¹ and in plants grown in a glasshouse during mid-summer (alternating shade and sun 300–1600 μmol m⁻² s⁻¹). To explain these differences between plants grown under low and high irradiances, Calvin cycle enzyme activities and metabolite content were determined. Activities of PRK and other non-equilibrium Calvin cycle enzymes fructose-1,6-bisphosphatase, sedoheptulose-1,7-bisphosphatase and ribulose-1,5-bisphosphate carboxylase-oxygenase were twofold higher in plants grown at 800 μmol m⁻² s⁻¹ or in the glasshouse than in plants grown at 330 μmol m⁻² s⁻¹. Activities of equilibrium enzymes transketolase, aldolase, ribulose-5-phosphate epimerase and isomerase were very similar under all growth irradiances. The flux control coefficient of 0.25 in plants grown at 330 μmol m⁻² s⁻¹ can be explained because low ribulose-5-phosphate content in combination with low PRK activity limits the synthesis of ribulose-1,5-bisphosphate. This limitation is overcome in high-light-grown plants because of the large relative increase in activities of sedoheptulose-1,7-bisphosphatase and fructose-1,6-bisphosphatase under these conditions, which facilitates the synthesis of larger amounts of ribulose-5-phosphate. This potential limitation will have maintained evolutionary selection pressure for high concentrations of PRK within the chloroplast. Received: 15 November 1999 / Accepted: 27 January 2000     

Salinity effects on CO2 assimilation and diffusive conductance of cowpea leaves

The effect of NaCl salinity at concentrations of 43–173 mM in nutrient solution on net gas exchange of attached cowpea [Vigna unguiculata (L.) Walp cv. California Black-eye No. 5 (CB5)] leaves was investigated under both greenhouse and growth chamber conditions.
There was a marked decrease in leaf conductance to water vapor after exposure to low salinity levels and a slighter decrease when salinity levels were higher. The decrease in net assimilation was much more gradual throughout the entire salinity range. The altered responses of net assimilation and leaf conductance to salinity were more evident at a high light intensity. A decrease in intercellular partial CO₂ pressure [p(CO₂)] was found at the low and intermediate salinity levels but not at the high level. These findings suggest that CO, assimilation was mainly controlled by stomatal conductance and the fixation of CO, might have been increased due to stimulated biochemical activity or to higher chlorophyll concentration per unit leaf area. A decrease in assimilation was already found one day after salinization and pro-ceeded up to 4 days when it was inhibited by 50% at 43 mM NaCl and up to 85% at 173 mM. The decrease in transpiration was larger than the decrease in net assimila-tion, and both were attributed to osmotic stress. Partial recovery was found thereaf-ter and new steady-state rates, in the range of 55 to 100% of the control, were then obtained for salinity levels between 43 and 130 mM. Inhibition of net CO, assimila-tion at this stage was attributed partly to a specific sodium effect and partly to plant water status. A linear relationship between leaf sodium content and net photosynthe-sis was also evident at this stage. Net CO, assimilation recovered more completely than transpiration when salt stress was removed, but at 173 mM NaCl recovery was neglible.     

Decrease in phosphoribulokinase activity by antisense RNA in transgenic tobacco. Relationship between photosynthesis, growth, and allocation at different nitrogen levels

To study the direct effects of photosynthesis on allocation of biomass by altering photosynthesis without altering leaf N or nitrate content, phosphoribulokinase (PRK) activity was decreased in transgenic tobacco (Nicotiana tabacum L.) with an inverted tobacco PRK cDNA and plants were grown at different N levels (0.4 and 5 mM NH4NO3). The activation state of PRK increased as the amount of enzyme was decreased genetically at both levels of N. At high N a 94% decrease in PRK activity had only a small effect (20%) on photosynthesis and growth. At low N a 94% decrease in PRK activity had a greater effect on leaf photosynthesis (decreased by up to 50%) and whole-plant photosynthesis (decreased by up to 35%) than at high N. These plants were up to 35% smaller than plants with higher PRK activities because they had less structural dry matter and less starch, which was decreased by 3- to 4-fold, but still accumulated to 24% to 31% of dry weight; young leaves contained more starch than older leaves in older plants. Leaves had a higher ion and water content, and specific leaf area was higher, but allocation between shoot and root was unaltered. In conclusion, low N in addition to a 94% decrease in PRK by antisense reduces the activity of PRK sufficient to diminish photosynthesis, which limits biomass production under conditions normally considered sink limited.     

Influence of CO2 on Reduction of Nitrate in Corn Seedlings

The influence of CO₂ on the assimilation of nitrate in intact corn seedlings was measured with ¹⁵N labelled nitrate, 24 and 48 h after the dark-grown seedlings were transferred to the light, either in normal air or in CO₂-free air. During the first 24 h CO₂ had no influence on nitrate reduction in intact seedlings. Experiments with detopped seedlings showed that during this period the roots were the only site of nitrate reduction. After 48 h seedlings grown in normal air had reduced more nitrate than detopped seedlings, and seedlings grown in CO₂-free air had reduced the same amount of nitrate as detopped seedlings. During the whole 48 h period CO₂ had no influence on the level of nitrate reductase of the leaves. It was concluded that in normal air corn leaves started to reduce nitrate after a lag period of 24 h and that in CO₂-free air they were incapable of nitrate reduction.     

The effect of growth at elevated CO2 concentrations on photosynthesis in wheat

Rising levels of atmospheric CO₂ will have profound, direct effects on plant carbon metabolism. In this study we used gas exchange measurements, models describing the instantaneous response of leaf net CO₂ assimilation rate (A) to intercellular CO₂ partial pressure (C_i), in vitro enzyme activity assay, and carbohydrate assay in order to investigate the photosynthetic responses of wheat (Triticum aestivum L., cv. Wembley) to growth under elevated partial pressures of atmospheric CO₂ (C_a). At flag leaf ligule emergence, the modelled, in vivo, maximum carboxylation velocity for RuBisCO was significantly lower in plants grown at elevated C_a than in plants grown at ambient C_a (70 Pa compared with 40 Pa). By 12 d after ligule emergence, no significant difference in this parameter was detectable. At ligule emergence, plants grown at elevated C_a exhibited reduced in vitro initial activities and activation states of RuBisCO. At their respective growth C_i values, the photosynthesis of 40-Pa-grown plants was sensitive to p(O₂) and to p(CO₂) whereas that of 70-Pa-grown plants was insensitive. Both sucrose and starch accumulated more rapidly in the leaves of plants grown at 70 Pa. At flag leaf ligule emergence, modelled non-photorespiratory respiration in the light (R_d) was significantly higher in 70-Pa-grown plants than in 40-Pa-grown plants. By 12 d after ligule emergence no significant differences in R_d were detectable.     

CO2 and intracellular pH

Abstract The experimental determination of cytoplasmic and vacuolar pH values is discussed. Despite variation in these values evidence indicates that intracellular pH values are normally regulated within narrow limits. The regulatory mechanisms proposed involve the metabolic consumption of OH^& and the active efflux of H ⁺. The evidence for intracellular pH modification in response to CO₂ hydration and the production of HCO^?₃ and H⁺ is examined. Theoretical calculations and experimental data indicate that CO₂ concentrations as high as 5% will lower intracellular pH. Conversely, variation in CO₂ levels around atmospheric concentrations is unlikely to perturb intracellular pH. High CO₂ levels are found in bulky tissues, and flooded root systems. Evidence is presented that the slow diffusion of dissolved CO₂ compared to gaseous CO₂ results in its accumulation. It is proposed that the accumulation of respiratory CO₂ may reduce intracellular pH values when plant tissues, cells or protoplasts are maintained in a liquid culture medium. Finally, the possible role of dark CO₂ fixation and organic acid synthesis in the regulation of intracellular pH is examined.     

Effects of photoinhibitory treatment on CO2 assimilation, the quantum yield of CO2 assimilation, D1 protein, ascorbate, glutathione and xanthophyll contents and the electron transport rate in vine leaves

The responses of photosynthesis to high light and low temperature were studied in vines cultivated in the greenhouse in low light. Exposure to high light (1000 /umol m^?2 s^?1) or low temperature (5 °C) alone had no measurable effect on the photosynthetic processes, but the combination of high light and low temperature caused rapid loss of photosynthetic capacity and a decrease in the efficiency of photosynthetic energy conversion. After a 15 h exposure to 5°C at high light, the F_v/sb/F_mratio had decreased by 80% and the apparent quantum yield by 75%. Nevertheless, when the leaves were returned to low light at 22°C, these parameters recovered rapidly. The foliar pools of ascorbate and glutathione decreased in the first hours of photoinhibitory treatment while the zeaxanthin content increased from negligible levels to about 50% of the total foliar xanthophyll pool. There was a clear correlation between the zeaxanthin content of the leaves and their F_v/F_m ratio during both photoinhibition and recovery. However, there was also a good correlation between the decrease in theF_v F_m ratio and the measured decrease in the total foliar levels of the antioxidants ascorbate and glutathione. The amount of D, protein diminished over the same period as the zeaxanthin levels were increasing. This approach, involving simultaneous measurements of several parameters considered to influence photosystemy II activity, clearly demonstrates that measured decreases in F_v/F_m may not simply be related to zeaxanthin levels or to amounts of D₁ protein alone but result from multifactoral influences.     

Estimation of the carbon dioxide (CO2) fertilization effect using growth rate anomalies of CO2 and crop yields since 1961

The effect of elevated carbon dioxide (CO₂) on crop yields is one of the most uncertain and influential parameters in models used to assess climate change impacts and adaptations. A primary reason for this uncertainty is the limited availability of experimental data on CO₂ responses for crops grown under typical field conditions. However, because of historical variations in CO₂, each year farmers throughout the world perform uncontrolled yield ‘experiments’ under different levels of CO₂. In this study, measurements of atmospheric CO₂ growth rates and crop yields for individual countries since 1961 were compared to empirically determine the average effect of a 1 ppm increase of CO₂ on yields of rice, wheat, and maize. Because the gradual increase in CO₂ is highly correlated with major changes in technology, management, and other yield controlling factors, we focused on first differences of CO₂ and yield time series. Estimates of CO₂ responses obtained from this approach were highly uncertain, reflecting the relatively small importance of year‐to‐year CO₂ changes for yield variability. Combining estimates from the top 20 countries for each crop resulted in estimates with substantially less uncertainty than from any individual country. The results indicate that while current datasets cannot reliably constrain estimates beyond previous experimental studies, an empirical approach supported by large amounts of data may provide a potentially valuable and independent assessment of this critical model parameter. For example, analysis of reliable yield records from hundreds of individual, independent locations (as opposed to national scale yield records with poorly defined errors) may result in empirical estimates with useful levels of uncertainty to complement estimates from experimental studies.     

CO2 assimilation, respiration and chlorophyll fluorescence in peach leaves infected by Taphrina deformans

Photosynthesis, respiration and chlorophyll fluorescence parameters were determined in peach ( Prunus persica L. cv. Dixired) leaves naturally infected by Taphrina deformans (Berk.) Tul. and in healthy leaves (controls), in two successive springs. A drastic decrease in net photosynthesis and an evident increase in respiration in curled leaves were noted. The instantaneous PSII fluorescence yield, with no (F₀) and with (F₀) quenching component, and steady state fluorescence yield (under actinic light, F_s) were essentially unchanged. Maximal fluorescence in dark-adapted (F_m) and illuminated (F'_m) leaves and the corresponding variable fluorescence (F_v and F_v) clearly decreased. The indicators of PSII quantum yield (F_v/F_m) in dark-adapted leaves, and the potential PSII excitation capture efficiency (F'_v/F'_m) and the quantum yield of PSII (q_p [F'_v/F'_m]) in the light were also significantly lower in curled leaves. Decreasing tendencies were also noted for the PSII photochemical yield (photochemical quenching, q_p) and in the energy status of the chloroplast (non-photochemical quenching, q_N, and Stern-Vollmer value, NPQ) although the differences were not always significant. In curled leaves the main alteration documented is the imbalance between the drastic inhibition of CO₂ fixation and the moderate decrease in photochemical reactions (i.e. F_v/F_m and ΔF/F'_m), indicating changes in the energy flux.     

Interannual climatic variation mediates elevated CO2 and O3 effects on forest growth

We analyzed growth data from model aspen (Populus tremuloides Michx.) forest ecosystems grown in elevated atmospheric carbon dioxide ([CO₂]; 518 μL L^?1) and ozone concentrations ([O₃]; 1.5 × background of 30–40 nL L^?1 during daylight hours) for 7 years using free‐air CO₂ enrichment technology to determine how interannual variability in present‐day climate might affect growth responses to either gas. We also tested whether growth effects of those gasses were sustained over time. Elevated [CO₂] increased tree heights, diameters, and main stem volumes by 11%, 16%, and 20%, respectively, whereas elevated ozone [O₃] decreased them by 11%, 8%, and 29%, respectively. Responses similar to these were found for stand volume and basal area. There were no growth responses to the combination of elevated [CO₂+O₃]. The elevated [CO₂] growth stimulation was found to be decreasing, but relative growth rates varied considerably from year to year. Neither the variation in annual relative growth rates nor the apparent decline in CO₂ growth response could be explained in terms of nitrogen or water limitations. Instead, growth responses to elevated [CO₂] and [O₃] interacted strongly with present‐day interannual variability in climatic conditions. The amount of photosynthetically active radiation and temperature during specific times of the year coinciding with growth phenology explained 20–63% of the annual variation in growth response to elevated [CO₂] and [O₃]. Years with higher photosynthetic photon flux (PPF) during the month of July resulted in more positive growth responses to elevated [CO₂] and more negative growth responses to elevated [O₃]. Mean daily temperatures during the month of October affected growth in a similar fashion the following year. These results indicate that a several‐year trend of increasingly cloudy summers and cool autumns were responsible for the decrease in CO₂ growth response.