Transient peaks in the delayed luminescence from Scenedesmus obtusiusculus induced by phosphorus starvation and carbon dioxide deficiency

Cells of the unicellular green alga Scenedesmus obtusiusculus (Chod.) were starved of phosphorus for 24, 48, 72 and 96 h, and the decay kinetics of the delayed luminescence from the differently starved cells was monitored for several minutes. Cells starved for 24 h showed similar delayed luminescence decay kinetics and accumulated output of photons as control cells after excitation with white light. Two transient peaks (with several components) in the decay kinetics of delayed luminescence were observed after 48 h of phosphorus starvation but not after 72 or 96 h. The amplitude of the transient peaks varied depending on the length of the excitation period with white light and on the length of the dark period preceding light excitation. High CO₂ availability induced no transient peak, whereas low CO₂ availability induced a high transient peak. Transient peaks could not be induced by excitation with light of 660 or 680 nm and only a single transient peak developed using 700 nm light. The kinetics of the delayed luminescence was changed, and the accumulated output of photons was decreased when the pH of the medium was changed from 7.2 to 9.5, both in cells starved for phosphorus for 96 h and in controls. The data indicate that a complicated metabolic pattern is involved in the mechanisms giving rise to the observed transient peaks in the delayed luminescence. The main factors may be a reduction in the translocation of trioses from chloroplasts, a concomitant reduction in Calvin cycle activities and changes in the amount of ATP and reducing agents available.     

Heterotrophic (Dark) CO2 Fixation by Euglena gracilis. Possible regulation by tricarboxylic acid cycle intermediates

SYNOPSIS. Heterotrophic (dark) CO₂ fixation by Euglena gracilis strain Z varies with phase of batch culture and mode of nutrition. Dark CO₂ fixation increased transiently during the growth of cells under photoautotrophic (CO₂, light) and heterotrophic (glucose, dark) conditions. Cells grown heterotrophically with acetate or ethanol had no transient increase in fixation. The addition of acetate to a heterotrophically growing culture during the period of increasing dark CO₂ fixation resulted in rapid elimination of this fixation. The results suggest that dark CO₂ fixation in Euglena functions in anaplerotic feeding of the tricarboxylic acid cycle, drained by biosyntheses during growth. Induction of the glyoxylate cycle by acetate may provide an alternate source of tricarboxylic cycle intermediates, obviating the requirement for dark CO₂ fixation as a source of the intermediates.     

Volume changes of Commelina communis guard cell protoplasts in response to K+, light and CO2

The effects of K⁺ concentration, light intensity and CO₂ levels on the volume of Commelina communis L. guard cell protoplasts were studied. Two degrees of swelling response were observed, both dependent on an external supply of K⁺, but not necessarily on the supply of a permeant anion. The presence of K⁺ itself, independent of light or CO₂ level, stimulated swelling at a relatively slow rate. When K⁺, light and low CO₂ conditions were supplied together, the swelling was relatively rapid and of high magnitude. The rapid swelling was specific for K⁺ and Rb⁺ giving a half maximal effect after 2 h at a KCl concentration of about 18 mmol m⁻³. The addition of CaCl₂ at 1 mol m⁻³ inhibited K⁺-dependent swelling under all conditions tested. The response to light and low CO₂ levels by Commelina guard cell protoplasts is thought to reflect a high degree of physiological integrity.     

Soil CO2 efflux in a boreal pine forest under atmospheric CO2 enrichment and air warming

The response of forest soil CO₂ efflux to the elevation of two climatic factors, the atmospheric concentration of CO₂ (↑CO₂ of 700 μmol mol⁻¹) and air temperature (↑ T with average annual increase of 5°C), and their combination (↑CO₂+↑ T ) was investigated in a 4-year, full-factorial field experiment consisting of closed chambers built around 20-year-old Scots pines ( Pinus sylvestris L.) in the boreal zone of Finland. Mean soil CO₂ efflux in May–October increased with elevated CO₂ by 23–37%, with elevated temperature by 27–43%, and with the combined treatment by 35–59%. Temperature elevation was a significant factor in the combined 4-year efflux data, whereas the effect of elevated CO₂ was not as evident. Elevated temperature had the most pronounced impact early and late in the season, while the influence of elevated CO₂ alone was especially notable late in the season. Needle area was found to be a significant predictor of soil CO₂ efflux, particularly in August, a month of high root growth, thus supporting the assumption of a close link between whole-tree physiology and soil CO₂ emissions. The decrease in the temperature sensitivity of soil CO₂ efflux observed in the elevated temperature treatments in the second year nevertheless suggests the existence of soil response mechanisms that may be independent of the assimilating component of the forest ecosystem. In conclusion, elevated atmospheric CO₂ and air temperature consistently increased forest soil CO₂ efflux over the 4-year period, their combined effect being additive, with no apparent interaction.     

The induction of the CO2 concentrating mechanism in a starch-less mutant of Chlamydomonas reinhardtii

In Chlamydomonas reinhardtii the formation of a starch sheath surrounding the pyrenoid is observed when cells grown under high CO₂ (5% CO₂ in air) are transferred to low CO₂ (0.03%) conditions. Formation of the starch sheath occurs coincidentally with induction of the CO₂ concentrating mechanism and with de novo synthesis of 5 polypeptides with molecular masses of 21, 36, 37, 42–44 kDa. We studied the effect of CO₂ concentrations on photosynthesis, ultrastructure and protein synthesis in Chlamydomonas reinhardtii cw-15 (wild phenotype for photosynthesis) and in the starch-less mutant BAFJ -6, with the aim to clarify the role of the pyrenoid starch sheath in the operation of the CO₂ concentrating mechanism and whether these low CO₂-inducible polypeptides are involved in the formation of starch sheath. When wild type and starch-less mutant cells were transferred from high to low CO₂, the CO₂ requirement for half-maximal rates of photosynthesis decreased from 40 μM to 2 μM CO₂. ³⁵SO₄^2- labeling analyses showed that the starch-less mutant induced the 5 low CO₂-inducible polypeptides. These observations suggest that the starch-less mutant was able to induce a fully active CO₂ concentrating mechanism. Since the starch-less mutant did not form a pyrenoid starch sheath, we suggest that the starch sheath is not involved in the operation of the CO₂ concentrating mechanism and that none of these 5 low CO₂-inducible proteins is involved in the formation of the starch sheath in Chlamydomonas .     

What physiological acclimation supports increased growth at high CO2 conditions?

Chlamydomonas acidophila Negoro is a green algal species abundant in acidic waters (pH 2–3.5), in which inorganic carbon is present only as CO₂. Previous studies have shown that aeration with CO₂ increased its maximum growth rate, suggesting CO₂ limitation under natural conditions. To unravel the underlying physiological mechanisms at high CO₂ conditions that enables increased growth, several physiological characteristics from high- and low-CO₂-acclimated cells were studied: maximum quantum yield, photosynthetic O₂ evolution (P_max), affinity constant for CO₂ by photosynthesis (K_0.5,p), a CO₂-concentrating mechanism (CCM), cellular Rubisco content and the affinity constant of Rubisco for CO₂ (K_0.5,r). The results show that at high CO₂ concentrations, C. acidophila had a higher K_0.5,p, P_max, maximum quantum yield, switched off its CCM and had a lower Rubisco content than at low CO₂ conditions. In contrast, the K_0.5,r was comparable under high and low CO₂ conditions. It is calculated that the higher P_max can already explain the increased growth rate in a high CO₂ environment. From an ecophysiological point of view, the increased maximum growth rate at high CO₂ will likely not be realised in the field because of other population regulating factors and should be seen as an acclimation to CO₂ and not as proof for a CO₂ limitation.     

CO2-concentrating mechanisms in Egeria densa,a submersed aquatic plant

Egeria densa is an aquatic higher plant which has developed different mechanisms to deal with photosynthesis under conditions of low CO₂ availability. On the one hand it shows leaf pH-polarity, which has been proposed to be used for bicarbonate utilization. In this way, at high light intensities and low dissolved carbon concentration, this species generates a low pH at the adaxial leaf surface. This acidification shifts the equilibrium HCO₃^–/CO₂ towards CO₂, which enters the cell by passive diffusion. By this means, E. densa increases the concentration of CO₂ available for photosynthesis inside the cells, when this gas is limiting. On the other hand, under stress conditions resulting from high temperature and high light intensities, it shows a biochemical adaptation with the induction of a C₄-like mechanism but without Kranz anatomy. Transfer from low to high temperature and light conditions induces increased levels of phospho enol pyruvate carboxylase (PEPC, EC 4.1.1.31) and NADP-malic enzyme (NADP-ME, EC 1.1.1.40), both key enzymes participating in the Hatch-Slack cycle in plants with C₄ metabolism. Moreover, one PEPC isoform, whose synthesis is induced by high temperature and light, is phosphorylated in the light, and changes in kinetic and regulatory properties are correlated with changes in the phosphorylation state of this enzyme. In the present review, we describe these two processes in this submersed angiosperm that appear to help it perform photosynthesis under conditions of extreme temperatures and high light intensities.     

Assimilate relations in source and sink leaves during acclimation to a CO2-enriched atmosphere

Evidence from previous studies suggested that adjustments in assimilate formation and partitioning in leaves might occur over time when plants are exposed to enriched atmospheric CO₂. We examined assimilate relations of source (primary unifoliolate) and developing sink (second mainstem trifoliolate) leaves of soybean [ Glycine max (L.) Merr. cv. Lee] plants for 12 days after transfer from a control (350 μl l⁻¹) to a high (700 μ l⁻¹) CO₂ environment. Similar responses were evident in the two leaf types. Net CO₂ exchange rate (CER) immediately increased and remained elevated in high CO₂. Initially, the additional assimilate at high CO₂ levels in the light and was utilized in the subsequent dark period. After approximately 7 days, assimilate export in the light began to increase and by 12 days reached rates 3 to 5 times that of the control. In the developing sink leaf, high rates of export in the light occurred as the leaf approached full expansion. The results indicate that a specific acclimation process occurs in source leaves which increases the capacity for assimilate export in the light phase of the diurnal cycle as plants adjust to enriched CO₂ and a more rapid growth rate.     

The effect of different growth conditions on dark and light carbon assimilation in Littorella uniflora

The effect of long-term exposure to different inorganic carbon, nutrient and light regimes on CAM activity and photosynthetic performance in the submerged aquatic plant, Littorella uniflora (L.) Aschers was investigated. The potential CAM activity of Littorella was highly plastic and was reduced upon exposure to low light intensities (43 μmol m⁻² s⁻¹), high CO₂ concentrations (5.5 mM, pH 6.0) or low levels of inorganic nutrients, which caused a 25–80% decline in the potential maximum CAM activity relative to the activity in the control experiments (light: 450 μmol m⁻² s⁻¹; free CO₂: 1.5 mM). The CAM activity was regulated more by light than by CO₂, while nutrient levels only affected the activity to a minor extent. The minor effect of low nutrient regimes may be due to a general adaptation of isoetid species to low nutrient levels.
The photosynthetic capacity and CO₂ affinity was unaffected or increased by exposure to low CO₂, irrespective of nutrient levels. High CO₂, low nutrient and low light, however, reduced the capacity by 22–40% and the CO₂ affinity by 35-45%, relative to control.
The parallel effect of growth conditions on CAM activity and photosynthetic performance of Littorella suggest that light and dark carbon assimilation are interrelated and constitute an integrated part of the carbon assimilation physiology of the plant. The results are consistent with the hypothesis that CAM is a carbon-conserving mechanism in certain aquatic plants. The investment in the CAM enzyme system is beneficial to the plants during growth at high light and low CO₂ conditions.     

The influence of elevated CO2 on species phenology, growth and reproduction in a Mediterranean old-field community

We studied the effects on the phenology, growth and reproduction of 19 Mediterranean species, of elevating the atmospheric CO₂ concentration ([CO₂]) to twice-ambient. Intact monoliths were taken from an old-field and put, during a six month growing season, into growth chambers in which external climatic conditions were mimicked and [CO₂] was regulated. Fruit set time was significantly changed in six species under elevated [CO₂] and leaf and branch senescence accelerated in most species. Grasses had fewer leaves and legumes were more branched at peak production under elevated [CO₂] than under ambient. Plant seed number was not significantly changed under elevated [CO₂], whereas the reproductive effort of grasses was significantly depressed. Reproductive and vegetative characteristics showed related responses to [CO₂], as species with enhanced biomass had a hastened fruit set time, a higher number of fruits per plant and a higher reproductive biomass under elevated [CO₂] than under ambient conditions, while species with depressed biomass had a delayed fruit set time, a lower number of fruits per plant and a lower reproductive biomass. Our results also show a high interspecific variability in [CO₂] response, but some trends emerged at the family level: the production of vegetative and reproductive modules were depressed in grasses and slightly stimulated in legumes.     

Light period of Crassulacean Acid Metabolism: Control of carbon transfer from malic acid to carbohydrates by CO2 concentration

In the CAM plants, Kalanchoë tubiflora (Harvey) Hasset, Sedum morganianum E. Walth and Sedum rubrotinctum R. T. Clausen, the effects of CO₂ concentrations on the light-dependent ¹⁴C transfer from the nocturnally synthetized [14C]-malic acid to starch have been studied. CO₂ concentrations up to 5 × 103 μ1 1–1 did not inhibit this carbon transfer. Higher CO₂ concentrations, however, were increasingly inhibitory. At 104 μl 1–1 CO₂, the carbon transfer was practically prevented.
The malic acid consumption in the light showed the same response to CO₂ concentrations as the [^l4C]-transfer. Photosynthesis itself was not inhibited by the CO₂ concentrations applied. It is assumed that, during phase III of CAM, light controls the internal CO₂ concentration via photosynthesis; and that the internal CO2 concentration then controls the rate of malate decarboxylation.     

The stomatal response to CO2 is linked to changes in guard cell zeaxanthin*

The mechanisms mediating CO₂ sensing and light–CO₂ interactions in guard cells are unknown. In growth chamber-grown Vicia faba leaves kept under constant light (500 μ mol m^–2 s^–1) and temperature, guard cell zeaxanthin content tracked ambient [CO₂] and stomatal apertures. Increases in [CO₂] from 400 to 1200 cm³ m^–3 decreased zeaxanthin content from 180 to 80 mmol mol^–1 Chl and decreased stomatal apertures by 7·0 μ m. Changes in zeaxanthin and aperture were reversed when [CO₂] was lowered. Guard cell zeaxanthin content was linearly correlated with stomatal apertures. In the dark, the CO₂-induced changes in stomatal aperture were much smaller, and guard cell zeaxanthin content did not change with chamber [CO₂]. Guard cell zeaxanthin also tracked [CO₂] and stomatal aperture in illuminated stomata from epidermal peels. Dithiothreitol (DTT), an inhibitor of zeaxanthin formation, eliminated CO₂-induced zeaxanthin changes in guard cells from illuminated epidermal peels and reduced the stomatal CO₂ response to the level observed in the dark. These data suggest that CO₂-dependent changes in the zeaxanthin content of guard cells could modulate CO₂-dependent changes of stomatal apertures in the light while a zeaxanthin-independent CO₂ sensing mechanism would modulate the CO₂ response in the dark.     

Development of Vacuoles and Vacuolar H+-ATPase Activity under Extremely High CO2 Conditions in Chiorococcum littorale Cells

Abstract: The number and cross-sectional area of vacuoles in Chlorococcum littorale cells visualized with a differential interfer ence fluorescence microscope increased after their transfer from air to 40% CO₂. An immunological observation indicated that the level of subunit B of vacuolar H*ATPase also increased under 40% CO₂ conditions. The activity of nitrate-sensitive ATP-ase associated with the vacuolar membrane was 2–fold higher in 40% CO₂-_-grown cells than in air-grown cells. The effects of inhi bitors on the ATPase activity confirmed that these activities were derived from vacuolar-type H-ATPase. These results sug gest that vacuole development associated with that of vacuolar H⁺-ATPase occurred during the acclimatization of C. littorale cells to extremely high CO₂ conditions.     

Effect of darkness and CO2 starvation on NH4+ and NO3− assimilation in the unicellular alga Cyanidium caldarium

Cyanidium caldarium (Tilden) Geitler, a non-vacuolate unicellular alga, resuspended in medium flushed with air enriched with 5% CO₂, assimilated NH₄⁺ at high rates both in the light and in the dark. The assimilation of NO₃⁻, by contrast, was inhibited by 63% in the dark. In cell suspensions flushed with CO₂-free air, NH₄⁺ assimilation decreased with time both in the light and in the dark and ceased almost completely after 90 min. The addition of CO₂ completely restored the capacity of the alga to assimilate NH₄⁺. NO₃⁻ assimilation, by contrast, was 33% higher in the absence of CO₂ and was linear with time. It is suggested that NO₃⁻ and NH₄⁺ metabolism in C. caldarium are differently controlled in response to the light and carbon conditions of the cell.     

Growth and physiological responses of canola (Brassica napus) to UV-B and CO2 under controlled environment conditions

Few studies have investigated the interaction of ultraviolet (UV)-B radiation and CO₂ concentration on plants. We studied the combined effects of UV-B radiation and CO₂ concentration on canola ( Brassica napus cv. 46A65) under four growth conditions – ambient CO₂ with UV-B (control), elevated CO₂ with UV-B, ambient CO₂ without UV-B, and elevated CO₂ without UV-B – to determine whether the adverse effects of UV-B are mitigated by elevated CO₂. Elevated CO₂ significantly increased plant height and seed yield, whereas UV-B decreased them. Elevated CO₂ ameliorated the adverse effects of UV-B in plant height. UV-B did not affect the physical characteristics of leaf but CO₂ did. Certain flower and fruit characteristics were affected negatively by UV-B and positively by CO₂. UV-B did not affect net photosynthesis, transpiration and stomatal conductance but decreased water use efficiency (WUE). Elevated CO₂ significantly increased net photosynthesis and WUE. Neither UV-B nor CO₂ affected chlorophyll (Chl) fluorescence. UV-B significantly decreased Chl b and increased the ratio of Chl a / b . Elevated CO₂ decreased only the ratio of Chl a / b . UV-B significantly increased UV-absorbing compounds while CO₂ had no effect on them. Both UV-B and CO₂ significantly increased epicuticular wax content. Many significant relationships were found between morphological, physiological, and chemical parameters. This study showed that elevated CO₂ can partially ameliorate some of the adverse effects of UV-B radiation in B . napus .     

Water stress, carbon dioxide, and light effects on sucrosephosphate synthase activity in Phaseolus vulgaris

The characteristics of sucrose-phosphate synthase (SPS; EC 2.4.1.14) activity in leaves of Phaseolus vulgaris L. cv. Linden was studied in plants subjected to water stress and various CO₂ and light treatments. When water was withheld for 3 days causing mild water stress (–0.9 MPa), the activity of SPS measured in crude extracts was reduced ca 50%. The effect of water stress was most evident when the enzyme was assayed with saturating amounts of its substrates fructose 6-phosphate and UDP glucose. Placing a water-stressed plant in an atmosphere containing 1% CO₂ reversed the effect of water stress on SPS activity over 5 h even though the water stress was not relieved. Holding unstressed leaves in low CO₂ partial pressure reduced the extractable activity of SPS. After 1 h of low CO₂ treatment the effect of low CO₂ could be reversed by 20 min of 5% CO₂. However, after 24 h of low CO₂ treatment, less SPS activity was recovered by the 20 min treatment. The cytosolic protein synthesis inhibitor cycloheximide prevented the slow recovery of SPS activity, but did not affect the rapid recovery of SPS. We conclude that the effect of water stress on SPS activity was a consequence of the inhibition of photosynthesis caused by stomatal closure. Responses of Phaseolus vulgaris SPS to light were similar to the response to low CO₂ in that the effects were most pronounced under V_max assay conditions. This is the first report of this type of light response of SPS in a dicotyledonous species.     

Light and CO2 responses of photosynthesis and chlorophyll fluorescence characteristics in bean plants after simulated acid rain

The effects of simulated acid rain on some chlorophyll fluorescence characteristics and photosynthetic gas exchange at different light intensities and CO₂ concentrations of bean plants were investigated. Measurements were carried out 3, 5 and 24 h after spraying. The results showed that a single acid rain (pH 1.8) treatment of bean plants reduced gas exchange, the maximal carboxylating efficiency and photochemical quenching. This treatment led also to increased CO₂ compensation point and non-photochemical quenching and changed the shape of CO₂ and light curves of photosynthesis. Both stomatal and non-stomatal factors contributed to the decreased photosynthetic rate, but their proportion changed with time of recovery of the photosynthetic ampparatus. Three hours after the treatment, the stomatal factors predominated in photosynthesis reduction, while during the next experimental period (5–24 h), mainly non-stomatal factors determined the decreased photosynthetic rate. It is suggested that the effects observed in consequence of acid rain treatment could be due to an increased intracellular accumulation of H⁺ and harmful ions contained in the cocktail. This probably led to impaired membrane permeability, enhancement of stroma acidity, uncoupled electron transport and insufficient accumulation of ATP and NADPH, which affected carbon metabolism.     

Photorespiratory nitrogen cycle – A critical evaluation

The photorespiratory nitrogen cycle proposed by Keys et al. (Nature 275: 741–743, 1978) involved formation of glycine by transamination of glyoxylate in the peroxisomes utilizing glutamate. Subsequently, glycine is oxidized to ammonia, serine and CO₂ in the mitochondria. The ammonia is reassimilated via the GS/GOGAT pathway generating glutamate. In this article, experimental evidence which suggests the occurrence of alternative mechanisms of glycolate and serine synthesis as well as of CO₂ and ammonia evolution is discussed. The problem of utilization of NADH coupled to ATP synthesis during photosynthesis is still unresolved, which complicates the glycine oxidation reaction in light. Further, factors are presented that determine the availability of amino donors in the peroxisomes and of amino acids viz., glycine, serine and glutamate for the operation of the photorespiratory N cycle. Recent evidence regarding the role of formate arising out of the reaction of glyoxylate with H₂O₂ in the regulation of photosynthetic electron flow in the Hill reaction, as well as of photorespiratory substrates functioning as carbon sources for the citric acid cycle in the light or for export to the growing tissues, suggests that the role of photo-respiration in plant metabolism needs to be reexamined.     

Stomatal responses to CO2 during a diel Crassulacean acid metabolism cycle in Kalanchoe daigremontiana and Kalanchoe pinnata

To investigate the diurnal variation of stomatal sensitivity to CO₂, stomatal response to a 30 min pulse of low CO₂ was measured four times during a 24 h time-course in two Crassulacean acid metabolism (CAM) species Kalanchoe daigremontiana and Kalanchoe pinnata , which vary in the degree of succulence, and hence, expression and commitment to CAM. In both species, stomata opened in response to a reduction in p CO₂ in the dark and in the latter half of the light period, and thus in CAM species, chloroplast photosynthesis is not required for the stomatal response to low p CO₂. Stomata did not respond to a decreased p CO₂ in K. daigremontiana in the light when stomata were closed, even when the supply of internal CO₂ was experimentally reduced. We conclude that stomatal closure during phase III is not solely mediated by high internal p CO₂, and suggest that in CAM species the diurnal variability in the responsiveness of stomata to p CO₂ could be explained by hypothesizing the existence of a single CO₂ sensor which interacts with other signalling pathways. When not perturbed by low p CO₂, CO₂ assimilation rate and stomatal conductance were correlated both in the light and in the dark in both species.