Changes of the chlorophyll fluorescence induction kinetics of C3 and CAM plants during day/night cycles

The induction kinetics of the 680 nm chlorophyll fluorescence were measured on attached leaves of Kalanchoë daigremontiana R. Hamet et Perr. (CAM plant), Sedum telephium L. and Sedum spectabile Bor. (C₃ plant in spring, CAM plant in summer) and Raphanus sativus L. (C₃ plant) at three different times during a 12/12h day/night cycle. During the fluorescence transient the fluorescence intensity at the O, P and T-level (f_O, f_max, f_st,) was different for the plant species tested; this may be due to their different leaf structure, pigment composition and organization of their photosystems. The kinetics of the fluorescence induction depended on the time of preillumination or dark adaptation during the light/dark cycle but not on the type of primary CO₂ fixation mechanism (C₃ and CAM). For dark adapted leaves measured either at the end of the dark phase or after dark adaptation of plants taken from the light phase a higher P-level fluorescence, a higher variable fluorescence (P-O) and a larger complementary area were found than for leaves of plants taken directly from the light phase. This indicates the presence of largely oxidized photosystem 2 acceptor pools during darkness. During the light phase the fluorescence decline after the P-level was faster than during the dark phase; from this we conclude that the light adaptation of the photosynthetic apparatus (state 1→ state 2 transition, Δ pH) during the induction period proceeded faster in plants taken from the light phase than in plants taken from the dark phase.     

The effects of salinity,crassulacean acid metabolism and plant age on the carbon isotope composition of <Emphasis Type="Italic">Mesembryanthemum crystallinum</Emphasis> L., a halophytic C<Subscript>3</Subscript>-CAM species

The carbon isotope composition of the halophyte Mesembryanthemum crystallinum L. (Aizoaceae) changes when plants are exposed to environmental stress and when they shift from C₃ to crassulacean acid metabolism (CAM). We examined the coupling between carbon isotope composition and photosynthetic pathway by subjecting plants of different ages to salinity and humidity treatments. Whole shoot ¹³C values became less negative in plants that were exposed to 400 mM NaCl in the hydroponic solution. The isotopic change had two components: a direct NaCl effect that was greatest in plants still operating in the C₃ mode and decreased proportionally with increasing levels of dark fixation, and a second component related to the degree of CAM expression. Ignoring the presumably diffusion-related NaCl effect on carbon isotope ratios results in an overestimation of nocturnal CO₂ gain in comparison to an isotope versus nocturnal CO₂ gain calibration established previously for C₃ and CAM species grown under well-watered conditions. It is widely taken for granted that the shift to CAM in M. crystallinum is partially under developmental control and that CAM is inevitably expressed in mature plants. Plants, cultivated under non-saline conditions and high relative humidity (RH) for up to 63 days, maintained diel CO₂ gas-exchange patterns and ¹³C values typical of C₃ plants. However, a weak CAM gas-exchange pattern and an increase in ¹³C value were observed in non-salt-treated plants grown at reduced RH. These observations are consistent with environmental control rather than developmental control of the induction of CAM in mature M. crystallinum under non-saline conditions.     

Le vie fotosintetiche C3, C4 e CAM nel contesto ecologico

Abstract

Ecological aspects of C₃, C₄ and CAM photosynthetic pathways. - Three different photosynthetic CO₂ fixation pathways are known to occur in higher plants. However all three pathways ultimately depend on the Calvin-Benson cycle for carbon reduction. The oxygenase activity of RuBP carboxilase is responsible for photorespiratory CO₂ release. Both C₄ and CAM pathways behave as a CO₂ concentrating mechanism which prevent photorespiration. The CO₂-concentrating mechanism in C₄ plants is based on intracellular symplastic transport of C₄ dicarboxylic acids from mesophyll-cells to the adjacent bundle-sheath cells. On the contrary in CAM plants the CO₂-concentrating mechanism is based on the intracellular transport of malic acid into and out of the vacuole.

The C₄ photosynthetic pathway as compared to the C₃ pathway permits higher rates of CO₂ fixation in high light and high temperature environments at low costs in terms of water loss, given the stability of the photosynthetic apparatus under such conditions.

CAM is interpreted as an adaptation to arid environments because it enables carbon assimilation to take place at very low water costs during the night when the evaporative demand is low. Nevertheless many aquatic species of Isoetes and some relatives are CAM, suggesting the adaptive role of CAM to environments which become depleted in CO₂.

The photosynthetic carbon fixation pathway certainly contributes to the ecological success of plants in different environments. However the distribution of plants may also reflect their biological history. On the other hand plants with different photosynthetic pathways coexist in many communities and tend to share resources in time. In any case some generalizations are possible: C₄ plants enjoy an ecological advantage in hot, moist, high light regions while the majority of species in desert environments are C₃; CAM plants are more frequent in semiarid regions with seasonal rainfall, coastal fog deserts, and in epiphytic habitats in tropical rain forests.     

Water relations of individual leaf cells ofMesembryanthemum crystallinum plants grown at low and high salinity

Summary The effects of saline conditions on the water relations of cells in intact leaf tissue of the facultative CAM plantMesembryanthemum crystallinum were studied using the pressure probe technique. During a 12-hr light/dark regime a maximum in turgor pressure was recorded for the mesophyll cells of salttreated (CAM) plants at the beginning of the light period followed 6 hr later by a pressure maximum in the bladder cells of the upper epidermis. In contrast, the turgor pressure in the bladder cells of the lower epidermis remained constant during light/dark regime. Turgor pressure maxima were not observed in untreated (C₃) plants.This finding strongly supports the assumption that water movement during malate accumulation and degradation in salttreated plants occurs predominantly between the mesophyll cells and the bladder cells of the upper epidermis. The necessary calculations take differences in the compartment volumes and in the elastic moduli of the cell walls () of the bladder cells of the lower and upper epidermis into account.Measurements of the kinetics of water transport showed that the half-time of water exchange for the two sorts of bladder cells were nearly identical in CAM plants and in C₃ plants. The absolute values of the half-times increased by about 45% in salttreated plants (about 113 sec) compared to the control plants (78 sec). Simultaneously, the half-time of water exchange of the mesophyll cells increased by about 60% from 14 sec (untreated plants) to 22 sec (salt-exposed plants). The leaves of this plant are apparently able to closely maintain the time of propagation of short-term osmotic pressure changes over a large salinity range.A cumulative plot of the data measured on both C₃ and CAM plants showed that the differences between the values of the elastic moduli of bladder cells from the lower and from the upper epidermis are due to differences in volume and suggested that the intrinsic elastic properties of the differently located bladder cells of C₃ and CAM plants were identical.A cumulative plot of the hydraulic conductivity of the membrane obtained both on mesophyll and on bladder cells of salttreated and of untreated plantsvs. the individual turgor pressure yielded a relationship well-known from giant algal cells and some higher plant cells: The hydraulic conductivity increased at very low pressure, indicating that the water permeability properties of the membrane of the various cell types of C₃ and CAM plants are pressure dependent, but otherwise identical.The results suggest that a few fundamental physical relationships control the adaptation of the tissue cells to salinity.     

Studies on carbon flow in Crassulacean acid metabolism during the initial light period

In the Crassulacean acid metabolism (CAM) plants Kalanchoë tubiflora and Sedum morganianum a shift in the pathways occurs by which external CO₂ enters the metabolism during the initial light period (phase II of the diurnal CAM cycle). At the beginning of phase II, CO₂ is fixed mainly by the C₄ pathway; during late phase II, however, it is fixed mainly via the C₃ pathway. The C₃ pathway contributes to the phosphoenolpyruvate-carboxylase-mediated CO₂ fixation by the provision of three-carbon skeletons. Since the shift in the carbon-flow pathway is delayed after a CO₂-free night when malic-acid accumulation in the vacuoles is prevented, it is very likely that the amount of malic acid in the vacuole is integrated in the mechanism which controls CAM during the initial light period. A light-on signal at the beginning of phase II is not required to bring about the shifts in the carbon-flow pathways, as is shown by the reaction of plants to a prolonged dark period. A model of carbon flow during phase II is proposed.Abbreviations CAM Crassulacean acid metabolism - PEP-Case phosphoenolpyruvate carboxylase     

Is Sedum acre L. a CAM plant?

Summary Sedum acre L. collected from its natural stands south of Darmstadt (Germany) showed ¹³C values typical for C₃ plants. This suggests that in situ at the natural stand CO₂ was fixed mainly via the C₃ mode of photosynthesis rather than via the CAM mode. However, experimental water stress shifts the CO₂ exchange pattern from the C₃ type to CAM type. Simultaneously, a diurnal rhythm of malic acid oscillation, typical for CAM, and increase of PEP-carboxylase and malic enzyme activities developed. Hence, Sedum acre is obviously to be classified as a facultative CAM plant. Because of the temperature characteristics of CO₂ exchange in Sedum acre, in situ CO₂ should be harvested from the atmosphere mainly during the seasons where water stress situations capable of inducing CAM are unlikely to occur.     

A comparative immunocytochemical localization study of phosphoenolpyruvate carboxylase in leaves of higher plants

The intracellular localization of phosphoenolpyruvate (PEP) carboxylase in plants belonging to the C₄, Crassulacean acid metabolism (CAM) and C₃ types was invetigated using an immunocytochemical method with an immune serum raised against the sorghum leaf enzyme. The plants studied were sorghum, maize (C₄ type), kalanchoe (CAM type), french bean, and spinach (C₃ type). In the green leaves of C₄ plants, it was shown that the carboxylase was located in the mesophyll and stomatic cells, being largely cytosolic in the mesophyll cells. Similarly, in CAM plants, the enzyme was found mainly outside the chloroplasts. In contrast, in C₃ plants, the PEP carboxylase appeared to be distributed between the cytosol and the chloroplasts of foliar parenchyma. Examination of sections from etiolated leaves showed fluorescence emission from etioplasts and cytosol for the parenchyma of french bean as well as for the bundle sheath and mesophyll of sorghum leaves. This data indicated that during the greening process photoregulation and evolution of PEP carboxylase is dependent on the tissue and on the metabolic type of the plant considered.Abbreviations CAM Crassulacean acid metabolism - PEP phosphoenolpyruvate     

Acquisition and assimilation of gaseous ammonia as revealed by intracellular pH changes in leaves of higher plants

Atmospheric ammonia (NH₃) from various anthropogenic sources has become a serious problem for natural vegetation. Ammonia not only causes changes in plant nitrogen metabolism, but also affects the acid-base balance of plants. Using the pH-sensitive fluorescent dyes pyranine and esculin, cytosolic and vacuolar pH changes were measured in leaves of C₃ and C₄ plants exposed for brief periods to concentrations of NH₃ in air ranging from 1.33 to 8.29 mol NH₃ · mol^-1 gas (0.94–5.86 mg · m^-3). After a lag phase, uptake of NH₃ from air at a rate of 200 nmol NH₃ · m ^{- 2} leaf area · s^{- 1} into leaves of Zea mays L. increased pyranine fluorescence indicating cytosolic alkalinisation. The increase was much larger in the dark than in the light. In illuminated leaves of the C₃ plant Pelargonium zonale L. and the C₄ plants Z. mays and Amaranthus caudatus L., NH₃-dependent cytosolic alkalinisation was particularly pronounced when CO₂ was supplied at very low levels (16 or 20 mol CO₂ · mol^{- 1} gas, containing 210 mmol O₂ · mol^{- 1} gas). An increase in esculin fluorescence, which was smaller than that of pyranine, was indicative of trapping of some of the NH₃ in the vacuoles of leaves of Spinacia oleracea L. and Z. mays. Photosynthesis and transpiration remained unchanged during exposure of illuminated leaves to NH₃, yielding an influx of 200 nmol NH₃ · m^-2 leaf area · s^-1 for up to 30 min, the longest exposure time used. Both CO₂ and O₂ influenced the extent of cytosolic alkalinisation. At 500 mol CO₂ · mol^-1 gas the cytosolic alkalinisation was suppressed more than at 16 or 20 mol CO₂ · mol^-1 gas. The suppressing effect of CO₂ on the NH₃induced alkalinisation was larger in illuminated leaves of the C₄ plants Z. mays and A. caudatus than in leaves of the C₃ plant P. zonale. A reduction of the O₂ concentration from 210 to 10 mmol O₂ · mol ^-1 gas, which inhibits photorespiration, increased the NH₃induced cytosolic alkalinisation in C₃ plants. Suppression by CO₂ or O₂ of the alkaline pH shift caused by the dissolution and protonation of NH₃ in queous leaf compartments, and possibly by the production of organic compounds synthesised from atmospheric NH₃, indicates that NH₃ which enters leaves is rapidly assimilated if photosynthesis or photorespiration provide nitrogen acceptor molecules.This work was supported by the Biotechnology and Biological Sciences Research Council and the Deutsche Forschungsgemein-schaft within the framework of the research of Sonderforschun-gsbreich 251 of the University of Würzburg. We are grateful to Dr. B. Wollenweber (The Royal Veterinary and Agricultural University, Denmark) for discussions.     

Induction of crassulacean acid metabolism in Mesembryanthemum crystallinum increases reproductive success under conditions of drought and salinity stress

Summary Mesembryanthemum crystallinum L., an inducible crassulacean acid metabolism (CAM) plant, was grown for approximately 5 weeks following germination in well-watered, non-saline soil in a controlled-environment chamber. During this time, plants were characterized by C₃ photosynthetic carbon metabolism. After the initial 5 weeks, CAM was induced by a combination of high soil salinity and reduced soil water content. One group of plants was allowed to engage in CAM by being continuously exposed to normal CO₂-containing air (about 350–400 ppm). A second group of plants was deprived of ambient CO₂ each night (12 h dark period) until completion of their life cycle, thereby minimizing potential carbon gain via dark CO₂ fixation. The capacity to express CAM under conditions of drought and salinity stress markedly improved reproductive success: plants kept in normal CO₂-containing air produced about 10 times more seeds than plants kept in CO₂-free air during dark periods. Seeds from plants deprived of ambient CO₂ overnight had more negative ¹³C values than seeds from plants kept in normal air.     

Photon yield of O2 evolution and chlorophyll fluorescence characteristics at 77 K among vascular plants of diverse origins

Photon yields of oxygen evolution at saturating CO₂ were determined for 44 species of vascular plants, representing widely diverse taxa, habitats, life forms and growth conditions. The photonyield values on the basis of absorbed light ( ^a) were remarkably constant among plants possessing the same pathway of photosynthetic CO₂ fixation, provided the plants had not been subjected to environmental stress. The mean ^a value ±SE for 37 C₃ species was 0.106±0.001 O₂·photon^-1. The five C₄ species exhibited lower photon yields and greater variation than the C₃ species ( ^a=0.0692±0.004). The ^a values for the two Crassulaceanacid-metabolism species were similar to those of C₃ species. Leaf chlorophyll content had little influence on ^a over the range found in normal, healthy leaves. Chlorophyll fluorescence characteristics at 77 K were determined for the same leaves as used for the photon-yield measurements. Considerable variation in fluorescence emission both at 692 nm and at 734 nm, was found 1) among the different species; 2) between the upper and lower surfaces of the same leaves; and 3) between sun and shade leaves of the same species. By contrast, the ratio of variable to maximum fluorescence emission at 692 nm (F_v/F_M, 692) remained remarkably constant (The mean value for the C₃ species was 0.832±0.004). High-light treatments of shade leaves resulted in a reduction in both ^a and the F_v/F_M, 692 ratio. The extent of the reductions increased with time of exposure to bright light. A linear relationship was obtained when ^a was plotted against F_v/F_M, 692. The results show that determinations of the photon yield of O₂ evolution and the F_v/F_M, 692 ratio can serve as excellent quantitative measures of photoinhibition of overall photosynthetic energy-conversion system and of photochemistry of photosystem II, respectively. This is especially valuable in field work where it is often impossible to obtain appropriate controls.Abbreviations and symbols CAM Crassulacean acid metabolism - PFD photon flux density (photon fluence rate) - PSI, PSII photosystem I, II - F_o, F_M, F_v instantaneous, maximum, variable fluorescence emission - absorptance - ^a photon yield (absorbed light) - ⁱ photon yield (incident light) C.I.W.-D.P.B. Publication No. 923     

Patterns of phosphoenolpyruvate carboxylase activity and cytosolic pH during light activation and dark deactivation in C3 and C4 plants

The rate and extent of light activation of PEPC may be used as another criterion to distinguish C₃ and C₄ plants. Light stimulated phosphoenolypyruvate carboxylase (PEPC) in leaf discs of C₄ plants, the activity being three times greater than that in the dark but stimulation of PEPC was limited about 30% over the dark-control in C₃ species. The light activation of PEPC in leaves of C₃ plants was complete within 10 min, while maximum activation in C₄ plants required illumination for more than 20 min, indicating that the relative pace of PEPC activation was slower in C₄ plants than in C₃ plants. Similarly, the dark-deactivation of the enzyme was also slower in leaves of C₄ than in C₃ species. The extent of PEPC stimulation in the alkaline pH range indicated that the dark-adapted form of the C₄ enzyme is very sensitive to changes in pH. The pH of cytosol-enriched cell sap extracted from illuminated leaves of C₄ plants was more alkaline than that of dark-adapted leaves. The extent of such light-dependent alkalization of cell sap was three times higher in C₄ leaves than in C₃ plants. The course of light-induced alkalization and dark-acidification of cytosol-enriched cell sap was markedly similar to the pattern of light activation and dark-deactivation of PEPC in Alternanthera pungens, a C₄ plant. Our report provides preliminary evidence that the photoactivation of PEPC in C₄ plants may be mediated at least partially by the modulation of cytosolic pH.Abbreviations CAM Crassulacean acid metabolism - G-6-P glucose-6-phosphate - PMSF phenylmethylsulfonyl fluoride - PEPC phosphoenolpyruvate carboxylase - PEPC-PK phosphoenolpyruvate ca carboxylase-protein kinase     

Recycling of respiratory CO2 during Crassulacean acid metabolism: alleviation of photoinhibition in Pyrrosia piloselloides

The regulation of Crassulacean acid metabolism (CAM) in the fern Pyrrosia piloselloides (L.) Price was investigated in Singapore on two epiphytic populations acclimated to sun and shade conditions. The shade fronds were less succulent and had a higher chlorophyll content although the chlorophyll a:b ratio was lower and light compensation points and dark-respiration rates were reduced. Dawn-dusk variations in titratable acidity and carbohydrate pools were two to three times greater in fronds acclimated to high photosynthetically active radiation (PAR), although water deficits were also higher than in shade fronds. External and internal CO₂ supply to attached fronds of the fern was varied so as to regulate the magnitude of CAM activity. A significant proportion of titratable acidity was derived from the refixation of respiratory CO₂ (27% and 35% recycling for sun and shade populations, respectively), as measured directly under CO₂-free conditions. Starch was shown to be the storage carbodydrate for CAM in Pyrrosia, with a stoichiometric reduction of C₃-skeleton units in proportion to malic-acid accumulation. Measurements of photosynthetic O₂ evolution under saturating CO₂ were used to compare the light responses of sun and shade fronds for each CO₂ supply regime, and also following the imposition of a photoinhibitory PAR treatment (1600 mol·m^-2·s^-1 for 3 h). Apparent quantum yield declined following the high-PAR treatment for sun- and shade-adapted plants, although for sun fronds CAM activity derived from respiratory CO₂ prevented any further reduction in photosynthetic efficiency. Recycling of respiratory CO₂ by shade plants could only partly prevent photoinhibitory damage. These observations provide experimental evidence that respiratory CO₂ recycling, ubiquitous in CAM plants, may have developed so as to alleviate photoinhibition.Abbreviations and symbols CAM Crassulacean acid metabolism - F_M maximal photosystem II fluorescence - F_T terminal steady-state fluorescence - PAR photosynthetically active radiation, 400–700 nm - H⁺ (dawn-dusk) variation in titratable acidity     

Light-induced pH changes in leaves of C4 plants

Light-induced changes in the fluorescence of the pH-indicating dyes pyranine or 5-(and 6-)carboxy-2, 7-dichlorofluorescein (CDCF) which had been fed to leaves were examined to monitor cellular pH changes. After short-term feeding of pyranine (pK 7.3) to leaves of Amaranthus caudatus L., a NAD-malic-enzyme-type C₄ plant, vascular bundles and surrounding cells became fluorescent. Fluorescence emission from mesophyll cells required longer feeding times. In CO₂-free air, pyranine fluorescence increased much more on illumination after mesophyll cells had become fluorescent than when only the vascular bundles and the bundle sheath of Amaranthus leaves had been stained. After short feeding times and in the absence of actinic illumination, CO₂ decreased pyranine fluorescence very slowly in Amaranthus and rapidly in C₃ leaves. After prolonged feeding times, the extent of the light-dependent increase in pyranine fluorescence was several times greater in different C₄ plants than in C₃ species. The kinetics of the fluorescence changes were also remarkably different in C₃ and C₄ plants. Carbon dioxide (500 l · l^–1) suppressed the light-induced increase in pyranine fluorescence more in C₄ than in C₃ leaves. Light-dependent changes in light scattering, which are indicative of chloroplast energization, and in 410-nm transmission, which indicate chloroplast movement, differed kinetically from those of the changes in pyranine fluorescence. Available evidence indicated that light-dependent changes in pyranine fluorescence did not originate from the apoplast of leaf cells. Microscopic observation led to the conclusion that, after prolonged feeding times or prolonged incubation, changes in pyranine fluorescence emitted from C₄ leaves reflect pH changes mainly in the cytosol of mesophyll cells. A transient acidification reaction indicated by quenching of pyranine fluorescence in the dark-light transient and not observed in C₃ species is attributed to the carboxylation of phosphoenolpyruvate. After short feeding times and in the absence of actinic illumination, CO₂ (250 l l^–1) decreased pyranine fluorescence very slowly in Amaranthus and more rapidly in C₃ leaves. After prolonged feeding times, both the rate and the extent of CO₂-dependent quenching of pyranine fluorescence increased, but the increase was insufficient to indicate the presence of highly active carbonic anhydrase in the compartment from which pyranine fluorescence was emitted. In contrast to pyranine, CDCF (pK 4.8) did not increase but rather decreased its fluorescence on illumination of an Amaranthus leaf, indicating acidification of an acidic compartment, most probably the vacuole of green leaf cells. The pattern of the acidification reaction was similar in C₄ and C₃ leaves. The remarkably large extent of the light-dependent increase in pyranine fluorescence from leaves of C₄ species and its slow kinetics are proposed to be caused by an alkalization of the cytosol which in the absence of CO₂ is larger in the mesophyll than in the bundle sheath. It gives rise to deprotonation of dye originally located in the mesophyll and, in addition, of dye which diffuses from the bundle sheath into the mesophyll following a pH gradient. Implications of slow diffusional transport of pyranine and CO₂ between mesophyll and bundle-sheath cells and the fast metabolite transport required in C₄ photosynthesis are discussed.Abbreviations CDCF 5-(and 6-)carboxy-2,7-dichlorofluorescein - DHAP dihydroxyacetone phosphate - PGA 3-phosphoglycerate This work was supported by the Sonderforschungsbereiche 176 and 251 of the University of Würzburg and by the Gottfried-Wilhelm-Leibniz Program of the Deutsche Forschungsgemeinschaft. A.S.R. was the recipient of a fellowship of the Alexander-von-Humboldt Foundation. We are grateful to Mrs. S. Neimanis for cooperation.     

Properties of phosphoenolpyruvate carboxylase in rapidly prepared,desalted leaf extracts of the Crassulacean acid metabolism plant Mesembryanthemum crystallinum L.

Properties of phosphoenolpyruvate (PEP) carboxylase, obtained from leaves of Mesembryanthemum crystallinum L. performing Crassulacean acid metabolism (CAM), were determined at frequent time points during a 12-h light/12-h dark cycle. Leaf extracts were rapidly desalted and PEP carboxylase activity as a function of PEP concentration, malate concentration, and pH was measured within 2 min after homogenization of the tissue. Maximum velocity of PEP carboxylase was similar in the light and dark at pH 7.5 and pH 8.0. However, PEP carboxylase had as much as a 12-fold lower K _m for PEP and as much as a 20-fold higher K _i for malate during the dark than during the light periods, the magnitude of these differences being dependent on the assay pH. Assuming that enzyme properties immediately after isolation reflect the approximate state of the enzyme in vivo, these differences in enzyme properties reduce the potential for CO₂ fixation via PEP carboxylase in the light. A small decrease in cytoplasmic pH in the light would greatly magnify the above differences in day/night properties of PEP carboxylase, because the sensitivity of PEP carboxylase to inhibition by malate increased with decreasing pH. Properties of PEP carboxylase were also studied in plants exposed to short-term perturbations of the normal 12-h light/12-h dark cycle (e.g., prolonged light period, prolonged dark period). Under all light/dark regimes, there was a close correlation between change in properties of PEP carboxylase and changes of the tissue from acidification to deacidification, and vice versa. Changes in properties of PEP carboxylase were not merely light/dark phenomena because they were also observed in plants exposed to continuous light or dark. the data indicate that, during CAM, PEP carboxylase exists in two stages which differ in their capacity for net malate synthesis. The physiologically-active state is distinguished by a low K _m for PEP and a high K _i for malate and favors malate synthesis. The physiologically-inactive state has a high K _m for PEP and a low K _i for malate and exists during periods of deacidification and other periods lacking synthesis of malic acid.Abbreviations CAM Crassulacean acid metabolism - PEP phosphoenolpyruvate - PEPC PEP carboxylase - RuBP ribulose 1,5-bisphosphate - RH relative humidity     

Light-dependent pH changes in leaves of C4 plants Comparison of the pH response to carbon dioxide and oxygen with that of C3 plants

Cytosolic and vacuolar pH changes caused by illumination or a changed composition of the gas phase were monitored in leaves of the NAD malic-enzyme-type C₄ plant Amaranthus caudatus L. and the C₃ plant Vicia faba L. by recording changes in the fluorescence of pH-indicating dyes which had been fed to the leaves. Light-dependent cytosolic alkalization and vacuolar acidification were maximal in the mesophyll cells under high-fluence-rate illumination and in the absence of CO₂. Under the same conditions, measurements of light scattering and electrochromic absorption changes at 518 nm revealed maximum thylakoid energization. The results show an intimate relationship between the energization of the photosynthetic apparatus by light, an increase in cytosolic pH and a decrease in vacuolar pH. This was true for both the C₄ and the C₃ plant, although kinetics, extent and even direction of cytosolic pH changes differed considerably in these plants, reflecting the differences in photosynthetic carbon metabolism. Darkening produced rapid acidification in Vicia, but not in Amaranthus. Continued alkalization in Amaranthus is interpreted to be the result of the decarboxylation of a C₄ intermediate and the release of liberated CO₂. In the presence of CO₂, energy consumption by carbon reduction decreased thylakoid energization, cytosolic alkalization and vacuolar acidification. Under low-fluence-rate illumination, thylakoid energization and light-dependent cytosolic and vacuolar pH changes were decreased in CO₂-free air compared with thylakoid energization and pH changes in 1% oxygen/99% nitrogen not only in the C₃ plant, but also in Amaranthus. Since oxygenation of ribulose bisphosphate initiates energy-consuming photorespiratory reactions in 21% oxygen, but not in 1% oxygen, this shows that photorespiratory reactions are active not only in the C₃ but also in the C₄ plant in the absence of external CO₂. Photorespiratory conditions appeared to decrease energization not only in the chloroplasts, but also in the cytosol. This is indicated by decreased transfer of protons from the cytosol into the vacuole, a process which is energy-dependent.Abbreviations CDCF 5-(and 6-)carboxy-2,7-dichlorofluorescein - P₇₀₀ electron-donor pigment in the reaction center of photosystem I - RuBP ribulose-1,5-bisphosphate This work was supported, within the framework of the Sonderforschungsbereiche 176 and 251 of the University of Würzburg, by the Gottfried-Wilhelm-Leibniz Program of the Deutsche Forschungsgemeinschaft. A.S.R. was the recipient of a fellowship from the Alexander-von-Humboldt-Foundation. We are grateful to Mr. Carsten Werner and Mrs. Spidola Neimanis for cooperation.     

Photosynthetic Pathway Types in Rangeland Plant Species from Inner Mongolia,North China

Photosynthetic pathway types, based on ¹³C measurements, were determined for 125 species in 95 genera and 32 families growing in rangelands from Inner Mongolia. Of the total species, 4 species from 3 genera and 2 families had C₄ photosynthesis (2 species in Gramineae and 2 in Chenopodiaceae) and 118 species from 90 genera and 31 families had C₃ photosynthesis. The number of C₄ species differed significantly among four rangeland sites, 4 species in desert, 3 species in steppe, but no C₄ species were identified in meadow and dune. Six species [e.g. Agriophyllum arenarium Bieb., Bassia dasyphylla O. Kuntze, Saussurea japonica (Thunb.) DC.] earlier identified as C₄ species using the enzyme ratio method were found as C₃ species using the carbon isotope ratios (¹³C). Hence the enzyme ratio method for C₃ and C₄ identification may not always be reliable. The ¹³C values of 3 species of Crassulaceae, which had been considered as CAM species, differed remarkably [–25.79 for Sedum aizoon L., –24.42 for Osostachys fimbriatus (Turcz.) Berger, and –16.97 for O. malacophyllus (Pall.) Fisch], suggesting that the use of ¹³C method as a diagnosis for CAM photosynthetic pathway type may not always be reliable and supplementary measurements are needed.     

Light dependence of quantum yields of Photosystem II and CO2 fixation in C3 and C4 plants

The light dependence of quantum yields of Photosystem II (_II) and of CO₂ fixation were determined in C₃ and C₄ plants under atmospheric conditions where photorespiration was minimal. Calculations were made of the apparent quantum yield for CO₂ fixation by dividing the measured rate of photosynthesis by the absorbed light [A/I=_CO2 and of the true quantum yield by dividing the estimated true rate of photosynthesis by absorbed light [(A+R_l)/I_a=_CO2·], where R_L is the rate of respiration in the light. The dependence of the _II/_CO2 and _II/_CO2 ^* ratios on light intensity was then evaluated. In both C₃ and C₄ plants there was little change in the ratio of _II/_CO2 at light intensities equivalent to 10–100% of full sunlight, whereas there was a dramatic increase in the ratio at lower light intensities. Changes in the ratio of _II/_CO2 can occur because respiratory losses are not accounted for, due to changes in the partitioning of energy between photosystems or changes in the relationship between PS II activity and CO₂ fixation. The apparent decrease in efficiency of utilization of energy derived from PS II for CO₂ fixation under low light intensity may be due to respiratory loss of CO₂. Using dark respiration as an estimate of R_L, the calculated _II/_CO2 ^* ratio was nearly constant from full sunlight down to approx 5% of full sunlight, which suggests a strong linkage between the true rate of CO₂ fixation and PS II activity under varying light intensity. Measurements of photosynthesis rates and _II were made by illuminating upper versus lower leaf surfaces of representative C₃ and C₄ monocots and dicots. With the monocots, the rate of photosynthesis and the ratio of _II/_CO2 exhibited a very similar patterns with leaves illuminated from the adaxial versus the abaxial surface, which may be due to uniformity in anatomy and lack of differences in light acclimation between the two surfaces. With dicots, the abaxial surface had both lower rates of photosynthesis and lower _II values than the adaxial surface which may be due to differences in anatomy (spongy versus palisade mesophyll cells) and/or light acclimation between the two surfaces. However, in each species the response of _II/_CO2 to varying light intensity was similar between the two surfaces, indicating a comparable linkage between PS II activity and CO₂ fixation.Abbreviations A measured rate of CO₂ assimilation - A+R_L true rate of CO₂ assimilation; e - CO₂ estimate of electrons transported through PSII per CO₂ fixed by RuBP carboxylase - f fraction of light absorbed by Photosystem II - F'_m yield of PSII chlorophyll fluorescence due to a saturating flash of white light under steady-state photosynthesis - F_s variable yield of fluorescence under steady-state photosynthesis; PPFD-photosynthetic photon flux density - I_a absorbed PPFD - PS II Photosystem II - R_d rate of respiration in the dark - R_I rate of respiration in the light estimated from measurement of R_d or from analysis of quantum yields - apparent quantum yield of CO₂ assimilation under a given condition (A/absorbed PPFD) - true quantum yield of CO₂ assimilation under a given condition [(A+R_L)/(absorbed PPFD)] - quantum yield for photosynthetic O₂ evolution - electrons transported via PS II per quantum absorbed by PS II Supported by USDA Competitive Grant 90-37280-5706.     

Analysis of inhibition of photosynthesis due to water stress in the C3 species Hordeum vulgare and Vicia faba: Electron transport, CO2 fixation and carboxylation capacity

A C₃ monocot, Hordeum vulgare and C₃ dicot, Vicia faba, were studied to evaluate the mechanism of inhibition of photosynthesis due to water stress. The net rate of CO₂ fixation (A) and transpiration (E) were measured by gas exchange, while the true rate of O₂ evolution (J _O2) was calculated from chlorophyll fluorescence analysis through the stress cycle (10 to 11 days). With the development of water stress, the decrease in A was more pronounced than the decrease in J _O2 resulting in an increased ratio of Photosystem II activity per CO₂ fixed which is indicative of an increase in photorespiration due to a decrease in supply of CO₂ to Rubisco. Analyses of changes in the J _O2 A ratios versus that of CO₂ limited photosynthesis in well watered plants, and RuBP pool/RuBP binding sites on Rubisco and RuBP activity, indicate a decreased supply of CO₂ to Rubisco under both mild and severe stress is primarily responsible for the decrease in CO₂ fixation. In the early stages of stress, the decrease in C _i (intercellular CO₂) due to stomatal closure can account for the decrease in photosynthesis. Under more severe stress, CO₂ supply to Rubisco, calculated from analysis of electron flow and CO₂ exchange, continued to decrease. However, C _i, calculated from analysis of transpiration and CO₂ exchange, either remained constant or increased which may be due to either a decrease in mesophyll conductance or an overestimation of C _i by this method due to patchiness in conductance of CO₂ to the intercellular space. When plants were rewatered after photosynthesis had dropped to 10–30% of the original rate, both species showed near full recovery within two to four days.Abbreviations A- net CO₂ assimilation rate - A ^*- net CO₂ assimilation rate plus dark respiration - ATP- adenosine triphosphate - CABP- carboxyarabinitol 1,5-bisphosphate - C _a- ambient CO₂ concentration - C _c- CO₂ concentration in the chloroplast - C _i- intercellular CO₂ concentration - E- transpiration rate - g _m- mesophyll conductance - g _s- stomatal conductance - J _O2 true rate of O₂ evolution - LSD- least significant difference - PPFD- photosynthetic photon flux density - PS II- Photosystem II - R _n- dark respiration rate - Rubisco- ribulose 1,5-bisphosphate carboxylase/oxygenase - RuBP- ribulose 1,5-bisphosphate - RWC- relative water content - _c- rate of carboxylation - _o- rate of oxygenation - _PSII- quantum yield of Photosystem II - - CO₂ compensation point in the absence of R _n - - water potential     

In situ studies on crassulacean acid metabolism in Sedum acre L. and Sedum mite Gil

Summary CO₂ exchange, the diurnal variations in the levels of malic, citric and isocitric acid, and the labelling pattern after ¹⁴CO₂ fixation were measured in Sedum acre and Sedum mite growing in situ. As predicted from laboratory experiments, drought changed the gas exchange pattern from a C₃ type to a crassulacean acid metabolism (CAM) type. This shift correlated with the development of a diurnal rhythm in the malic acid content. The results of ¹⁴CO₂ pulse-chase experiments suggest that in well-watered plants a CAM pattern of carbon flow already exists; hence water stress might enhance latent CAM rather than induce it. The in situ CAM performance by the Sedum species appeared to be highly susceptible to modulation by season and external factors, particularly light and temperature.CAM did not substantially contribute to total carbon gain in S. acre and S. mite. During most of their lifecycles the plants grow under conditions that favour CO₂ uptake by the C₃ pathway rather than by CAM. Hence, despite a capability to feature CAM, the ¹³C values found in S. acre and S. mite are those of C₃ plants.Abbreviations CAM Crassulacean Acid Metabolism - PEP-C Phosphoenolpyruvate-Carboxylase - DW Dry weight Dedicated to Prof. Dr. Dr. h.c. M. Evenari on the occasion of his 75th birthday and to Dr. K.F. Springer     

Physiological and biochemical aspects and molecular mechanisms of plant adaptation to the elevated concentration of atmospheric CO<Subscript>2</Subscript>

The review of publications concerning the impact of increasing CO₂ concentration in the Earths atmosphere (C_a) on higher terrestrial plants. The physiological changes in plants induced by increasing C_a, including growth and biochemical composition, the characteristics of photosynthesis and respiration, as well as the molecular mechanisms of the regulation of the activity of most important biosynthetic enzymes at early and late stages of the exposure to elevated C_a are under consideration. Various concepts of metabolic regulation during acclimation to increasing CO₂ concentration are critically reviewed. The pathways of possible involvement of carbonic anhydrase-mediated systems of CO₂ transport and concentration during C₃ photosynthesis of higher plants, the metabolic and signal mechanisms of photosynthesis inhibition by carbohydrates and the role of ethylene at elevated C_a are presented. The effect of elevated C_a on plant development and source-sink relations, as well as its interaction with other environmental factors, such as mineral, primarily nitrogen nutrition, light, temperature, and water regime, are discussed in with the context of potential forecasting of the consequences of increase in C_a and temperature for the activities of various higher plant forms in the rapidly changing climate.Translated from Fiziologiya Rastenii, Vol. 52, No. 1, 2005, pp. 129–145.Original Russian Text Copyright © 2005 by Romanova.