菠萝叶片绿色组织与贮水组织的腺苷酸及焦磷酸水平的昼夜变化

研究了景天酸代谢(CAM)植物菠萝(Ananascomosus)叶片绿色组织与贮水组织的苹果酸、腺苷酸及焦磷酸含量的昼夜变化。夜间苹果酸的积累仅发生在绿色组织中,而且,其含量也远高于贮水组织。绿色组织中能荷和无机磷含量夜间增高,白天下降。绿色组织中焦磷酸含量夜间增加,在白天的头几个小时迅速下降到低的水平,然后保持稳定。与绿色组织相比,贮水组织中ATP、ADP、无机磷和焦磷酸的含量低得多,且不表现昼夜变化,在贮水组织中没有测到AMP。     

Day-night changes of citric-acid levels in crassulacean acid metabolism: phenomenon and ecophysiological significance

Abstract A reappraisal is offered of old and new observations of substantial day-night changes of citric-acid levels in crassulacean acid metabolism (CAM). In contrast to malic acid, the biochemical consequences and the ecophysiological significance of nocturnal accumulation of citric acid in CAM have not received due attention. Considerations show that citric-acid accumulation does not provide a means for nocturnal storage of CO₂. It may, however, serve carbon retention by internal recycling and sustain the water budget affording a vacuolar osmoticum. Since citric-acid accumulation energetically is considerably more favourable than malic-acid accumulation, this may have important ecophysiological implications. The questions raised by these reflections can and need to be tackled experimentally.     

菠萝叶片绿色组织与贮水组织液泡膜ATPase和焦磷酸酶特性

研究了景天酸代谢 (CAM)植物菠萝叶片绿色组织与贮水组织液泡膜ATPase和焦磷酸酶 (PPase)特性。绿色组织有较高的ATPase和PPase活力。贮水组织 (绿色组织 )ATPase和PPase活力的最佳温度分别为 43和 49℃ (3 7和 46℃ )。当温度小于最佳温度时 ,ATPase和PPase活力随着温度的上升而上升 ;而当温度大于最佳温度时 ,ATPase和PPase活力随着温度的上升而下降。在 (1 0～ 46℃ ) 1 0～3 7℃的温度范围 ,绿色组织 (PPase)ATPase活力随着温度的上升而上升的速度大于贮水组织中(PPase)ATPase活力上升的速度 ,绿色组织有 (相对 )较高的ATPase和PPase活力 ;但在 (49～ 6 1℃ )43～ 5 5℃的温度范围 ,贮水组织中 (PPase)ATPase活力随着温度的上升而下降的速度小于绿色组织中(PPase)ATPase活力下降的速度 ,贮水组织有相对较高的ATPase和PPase活力。此外 ,贮水组织中这两种酶活力的热稳定性强于绿色组织中这两种酶活力的热稳定性。对这两种酶活力的最佳 pH和底物依赖性也进行了研究     

菠萝叶片绿色组织与贮水组织中代谢物水平的昼夜变化

研究了景天酸代谢(CAM)植物菠萝叶片绿色组织与贮水组织(WSP)的苹果酸、柠檬酸、异柠檬酸、淀粉、果糖、葡萄糖、蔗糖、葡糖-1-磷酸(G-1-P)、葡糖-6-磷酸(G-6-P)、果糖-6-磷酸(F-6-P)、草酰乙酸(OAA)及磷酸烯醇式丙酮酸(PEP)水平的昼夜变化。夜间苹果酸的积累仅发生在绿色组织中,表明只有绿色组织才能进行CAM。可溶性已糖(葡萄糖和果糖)是绿色组织中夜间苹果酸累积的主要碳源。绿色组织G-1-P、G-6-P和F-6-P水平在夜间的初期上升,后期下降,昼间的头3h仍下降,3h后变化不明显。绿色组织中OAA和PEP水平也发生昼夜变化。在贮水组织中没有测到淀粉、蔗糖、OAA和PEP。除葡萄糖和果糖外,WSP中其它代谢物的含量都远低于绿色组织,而且WSP中所有代谢物都无明显的昼夜变化。     

Endogenous rhythms and chaos in crassulacean acid metabolism

Endogenous free-running regular circadian oscillations of net CO₂ exchange in the crassulacean-acidmetabolism (CAM) plant Kalanchoë daigremontiana Hamet et Perrier de la Bâthie under constant external conditions in continuous light have been shown to change to irregular non-predictable (chaotic) time behaviour as irradiance or temperature are raised above a critical level. A model of CAM has been constructed with pools of major metabolites of varying concentrations, flows of metabolites leading to exchange between pools, metabolite transformations determined by chemical reactions, and feedback regulations. The model is described by a system of coupled non-linear differential equations. It shows stable rhythmicity in normal dark-light cycles and in continuous light and, like the K. daigremontiana leaves in the experiments, a change to chaos as irradiance is increased. The maintenance of endogenous oscillations in the model is brought about by a hysteresis switch or beat oscillator between two stable oscillation modes. In CAM these stable modes are vacuolar malate accumulation and remobilization. The model shows that the physical nature of the beat oscillator in the leaves can be explained by the balance between active and passive transport at the tonoplast.Abbreviations CAM crassulacean acid metabolism - D dark period - DL 12:12 h dark-light rhythm - L light period - LL constant illumination - PPFD photosynthetic photon flux density - T^L leaf temperature It is a great pleasure to thank Dr. G.-H. Vieweg, (Roßdorf, FRG) for his long-lasting efforts to have the phytotron in Darmstadt erected and for his persistent involvement during the various phases of planning and building. This made the present experiments possible. Dr. D. Kramer is thanked for all the time he spends to maintain functioning of the facility. Dr. P. Keller and Ms. Erika Ball assisted with the gas-exchange technology and helped with the surveillance of the long-running experiments, and Ms. Erika Ball performed all the integrations. Ms. Doris Schäfer is thanked for drawing the gas-exchange curves for publication. We are also most grateful to Professor Chr. Giersch and Professor M. Kluge (both Institut für Botanik, Technische Hochschule Darmstadt, FRG) for valuable discussions.     

Day-night changes in the levels of adenine nucleotides,phosphoenolpyruvate and inorganic pyrophosphate in leaves of plants having Crassulacean acid metabolism

The levels of phosphorylated compounds studied during the dark period of Crassulacean acid metabolism (CAM) in Kalanchoë leaves showed increases for ATP and pyrophosphate and decreases for ADP, AMP and phosphenolpyruvate; levels of inorganic phosphate remained constant. Changes in adenylate levels and the correlated nocturnal increase in adenylate-energycharge were closely related to changes in malate levels. The increase in ATP levels was much inhibited in CO₂-free air and stimulated after induction of CAM in short-day-treated plants of K. blossfeldiana cv. Tom Thumb. Changes in levels of phosphoenolpyruvate and pyrophosphate were independent of the presence of CO₂. The results show the operation of complex regulatory mechanisms in the energy metabolism of CAM plants during nocturnal malic-acid accumulation.Abbreviations CAM Crassulacean acid metabolism - FW fresh weight - OAA oxaloacetic acia - PEP phosphoenol pyruvate - PP_i pyrophosphate     

Eddy covariance captures four‐phase crassulacean acid metabolism (CAM) gas exchange signature in Agave

Mass and energy fluxes were measured over a field of Agave tequilana in Mexico using eddy covariance (EC) methodology. Data were gathered over 252 d, including the transition from wet to dry periods. Net ecosystem exchanges (F_N,EC) displayed a crassulacean acid metabolism (CAM) rhythm that alternated from CO₂ sink at night to CO₂ source during the day, and partitioned canopy fluxes (F_A,EC) showed a characteristic four‐phase CO₂ exchange pattern. Results were cross‐validated against diel changes in titratable acidity, leaf‐unfurling rates, energy exchange fluxes and reported biomass yields. Projected carbon balance (g C m^?2 year^?1, mean ± 95% confidence interval) indicated the site was a net sink of ?333 ± 24, of which contributions from soil respiration were +692 ± 7, and F_A,EC was ?1025 ± 25. EC estimated biomass yield was 20.1 Mg (dry) ha^?1 year^?1. Average integrated daily F_A,EC was ?234 ± 5 mmol CO₂ m^?2 d^?1 and persisted almost unchanged after 70 d of drought conditions. Regression analyses were performed on the EC data to identify the best environmental predictors of F_A. Results suggest that the carbon acquisition strategy of Agave offers productivity and drought resilience advantages over conventional semi‐arid C3 and C4 bioenergy candidates.     

Latitudinal variation in the degree of crassulacean acid metabolism in Puya chilensis

Crassulacean acid metabolism (CAM) is a photosynthetic pathway found in many plant species from arid and semiarid environments. Few studies aiming to characterise plant species as CAM or C₃ account for inter‐population differences in photosynthetic pathway, often relying on samples taken from herbarium material and/or a single plant or population. This may be especially problematic for species growing under contrasting climate conditions, as is the case for species with a wide geographic range. We used Puya chilensis, a species previously reported as CAM and C₃, to study among‐population variation in expression of the CAM pathway within its distribution range, which spans a significant climate gradient. We carried out a wide sampling scheme, including five populations and a combination of analytical methods (quantification of nocturnal acidification and stable isotope measurements). The study populations of P. chilensis encompass the entire latitudinal distribution range, from semi‐arid to temperate oceanic climates. Our results indicate that CAM decreased with latitude. However, even in the southern (wetter) populations, where δ¹³C values were indicative of C₃ metabolism, we found some nocturnal acidification. We stress the value of using two methods along with the use of samples from different populations, as this allows more reliable conclusions on the photosynthetic pathway for ‘probable’ CAM species that face varying climate conditions within their distribution ranges.     

Peeling back the layers of crassulacean acid metabolism: functional differentiation between Kalanchoë fedtschenkoi epidermis and mesophyll proteomes

Crassulacean acid metabolism (CAM) is a specialized mode of photosynthesis that offers the potential to engineer improved water‐use efficiency (WUE) and drought resilience in C₃ plants while sustaining productivity in the hotter and drier climates that are predicted for much of the world. CAM species show an inverted pattern of stomatal opening and closing across the diel cycle, which conserves water and provides a means of maintaining growth in hot, water‐limited environments. Recent genome sequencing of the constitutive model CAM species Kalanchoë fedtschenkoi provides a platform for elucidating the ensemble of proteins that link photosynthetic metabolism with stomatal movement, and that protect CAM plants from harsh environmental conditions. We describe a large‐scale proteomics analysis to characterize and compare proteins, as well as diel changes in their abundance in guard cell‐enriched epidermis and mesophyll cells from leaves of K. fedtschenkoi. Proteins implicated in processes that encompass respiration, the transport of water and CO₂, stomatal regulation, and CAM biochemistry are highlighted and discussed. Diel rescheduling of guard cell starch turnover in K. fedtschenkoi compared with that observed in Arabidopsis is reported and tissue‐specific localization in the epidermis and mesophyll of isozymes implicated in starch and malate turnover are discussed in line with the contrasting roles for these metabolites within the CAM mesophyll and stomatal complex. These data reveal the proteins and the biological processes enriched in each layer and provide key information for studies aiming to adapt plants to hot and dry environments by modifying leaf physiology for improved plant sustainability.     

CAM activity and day/night changes in the ultrastructure of stem chlorenchyma of Cissus quadrangularis L. as influenced by drought

Abstract Stem chlorenchymatous cells from well-watered and water-stressed plants of Cissus quadrangularis L. were examined to evaluate the effect of drying on cell structure and metabolism. At a stem relative water content as low as 52-58%, cellular integrity was maintained, but some qualitative and quantitative alterations occurred. Chlorophyll coment was higher in stressed plants, and the intact photosynthetic apparatus exhibited an increased degree of thylakoid stacking paralleled by a lower chlorophyll a/h ratio. Overnight malate accumulation was very low. Starch was retained but its day/night fluctuation was practically suppressed. Plastosomes were smaller and fewer, and the range of their daily fluctuation was very reduced. Vacuoles underwent fragmentation and accumulated osmiophilic deposits. The maintenance of the cellular integrity accounts for the ability of C. quadrangularis to recover rapidly from water stress.     

Comparative studies on immunological and molecular properties of phosphoenolpyruvate carboxylase in species of Sedum and Kalanchoë performing crassulacean acid metabolism (CAM)

Abstract. PEP-carboxylase was extracted and partially purified from nine species of the genus Sedum and three species of the genus Kalanchoe, all performing CAM. Immunological and molecular properties of these enzymes were compared. Molecular weight estimation with gradient slab gels showed identical molecular weights of about 232,000 for all PEP-carboxylases. Ouchlerlony double-diffusion analysis, immunotitralion and SDS polyacrylamide clectrophoresis indicated the presence of PEP-c dimers consisting of monomers of MW 105,000 and 115,000. A model of PEP-c substructure is proposed. The results are discussed in the context of CAM performance in the genus Sedum     

Inducibility of crassulacean acid metabolism (CAM) in Clusia species; physiological/biochemical characterisation and intercellular localization of carboxylation and decarboxylation processes in three species which exhibit different degrees of CAM

The biochemical basis for photosynthetic plasticity in tropical trees of the genus Clusia was investigated in three species that were from contrasting habitats and showed marked differences in their capacity for crassulacean acid metabolism (CAM). Physiological, anatomical and biochemical measurements were used to relate changes in the activities/amounts of key enzymes of C₃ and C₄ carboxylation to physiological performance under severe drought stress. On the basis of gas-exchange measurements and day/night patterns of organic acid turnover, the species were categorised as weak CAM-inducible (C.aripoensis Britt.), C₃-CAM intermediate (C. minor L.) and constitutive CAM (C.␣rosea Jacq. 9.). The categories reflect genotypic differences in physiological response to drought stress in terms of net carbon gain; in C. aripoensis net carbon gain was reduced by over 80% in drought-stressed plants whilst carbon gain was relatively unaffected after 10 d without water in C. rosea. In turn, genotypic differences in the capacity for CAM appeared to be directly related to the capacities/amounts of phosphoenolpyruvate carboxylase (PEPCase) and phosphoenolpyruvate carboxykinase (PEPCK) which increased in response to drought in both young and mature leaves. Whilst measured activities of PEPCase and PEPCK in well-watered plants of the C₃-CAM intermediate C. minor were 5–10 times in excess of that required to support the magnitude of organic acid turnover induced by drought, close correlations were observed between malate accumulation/PEPCase capacity and citrate decarboxylation/PEPCK capacity in all the species. Drought stress did not affect the amount of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) protein in any of the species but Rubisco activity was reduced by 35% in the weak CAM-inducible C. aripoensis. Similar amounts of glycine decarboxylase (GDC) protein were present in all three species regardless of the magnitude of CAM expression. Thus, the constitutive CAM species C. rosea did not appear to show reduced activity of this key enzyme of the photorespiratory pathway, which, in turn, may be related to the low internal conductance to CO₂ in this succulent species. Immuno-histochemical techniques showed that PEPCase, PEPCK and Rubisco were present in cells of the palisade and spongy parenchyma in leaves of species performing CAM. However, in leaves from well-watered plants of C. aripoensis which only performed C₃ photosynthesis, PEPCK was localized around latex-producing ducts. Differences in leaf anatomy between the species suggest that the association between mesophyll succulence and the capacity for CAM in these hemi-epiphytic stranglers has been selected for in arid environments. Received: 4 July 1997 / Accepted: 27 November 1997     

Nocturnal citrate accumulation and its response to environmental stress in the CAM plant Kalanchoe pinnata (Lam.) Pers.

Abstract. Kalanchoe pinnata (Lam.) Pers., a plant having crassulacean acid metabolism (CAM), was grown in high light (16–23 mol photons m⁻² d⁻¹) and in the shade (0.8–2.1 mol photons m⁻² d⁻¹), respectively. Plants were stressed in three ways, i.e. by transfer from high light to shade or vice versa just before measurements, and by withholding nitrogen and/or water. During the day-night cycle of CAM, K. pinnata showed day-night changes of citrate levels (Δ citrate) in addition to malate changes (Δ malate). Changes of leaf-cell sap osmotic pressure. Δπ, were linearly correlated with these changes of organic-acid anion levels with a relation of Δπ/(Δ citrate +Δ malate) = 1/1. The environmental stressor, i.e. limited N-nutrition, drought and higher or lower irradiance than experienced during growth, affected the absolute and relative contributions made by Δ citrate and Δ malate to total nocturnal organic-acid accumulation. In the high-light-grown plants transferred to the shade, changes of citrate levels were much less affected than changes of malate levels by the generally decreased metabolic activity and inhibition of CO₂ uptake. In the shade-grown plants, Δ citrate increased in response to stress imposed by interactive effects of the three stressors.     

Water flow and water storage in Agave deserti: osmotic implications of crassulacean acid metabolism

Abstract Water flow and water storage were investigated for Agave deserti, a desert succulent showing crassulacean acid metabolism (CAM). The anatomy and water relations of the peripheral chlorenchyma, where CAM occurs, and the central water-storage parenchyma were investigated for its massive leaves so that these tissues could be incorporated as discrete elements into an electrical-circuit analogue of the whole plant. The daily cycling of osmotic pressure was represented by voltage sources in series with the storage capacitors. With soil water potential and leaf transpiration rate as input variables, axial water flow through the vascular bundles and radial flows into and out of storage during the day/night cycle were determined. The predominantly nocturnal transpiration was coincident with increases in cell osmotic pressure and in titratable acid of the leaf chlorenchyma. In the outer layers of the chlorenchyma, water potential was most negative at the beginning of the night when transpiration was maximum, while the water-storage parenchyma reached its minimal water potential 9 h later. The roots plus stem contributed 7% and the leaves contributed 50% to the total water flow during maximal transpiration; peak water flow from the soil to the roots occurred at dawn and was only 58% of the maximal transpiration rate. Over each 24-h period, 39% of the water lost from the plant was derived from storage, with flow into storage occurring mainly during the daytime. Simulations showed that the acid accumulation rhythm of CAM had little impact on water uptake from the soil under the conditions employed. In the outer chlorenchyma, water potential and water flows were more sensitive to the day/night changes in transpiration than in osmotic pressure. Nevertheless, cell osmotic pressure had a large influence on turgor pressure in this tissue and determined the extent to which storage was recharged during the latter part of the night.     

Oscillatory model of crassulacean acid metabolism: structural analysis and stability boundaries with a discrete hysteresis switch

We propose a simple oscillatory model of crassulacean acid metabolism (CAM) describing the CO₂ uptake and nocturnal acidification of CAM plants by a system of coupled non-linear differential equations. Large differences in the content of metabolite pools are treated using a pseudo-steady-state approach. For the first time, simulations of the CAM cycle investigate its dependence on all three major control parameters simultaneously: temperature, photon flux density and external CO₂ concentration. Under stationary conditions in time the model shows either endogenous rhythmicity or two distinct steady states. Stability boundaries are calculated in parameter space.     

The appearance of a new molecular species of phosphoenolpyruvate carboxylase (PEPC) and the rapid induction of CAM in Sedum telephium L.

Abstract. When detached leaves of Sedum telephium are incubated in the absence of water, a rapid switch from C₃ photosynthesis to CAM (as indicated by the onset of day-to-night fluctuations in titratable acidity. ΔH⁺) occurs within the first dark period. The C₃-CAM switch in intact plants occurs within 3 5d. Extractable activity of phospho enol pyruvate carboxylase (PEPC) increases five-fold in intact plants during CAM induction; however, during rapid CAM induction in detached leaves, there is only a very small increase in PEPC activity. Fractionation by anion exchange chromatography of crude extracts from leaves of intact plants subjected to water deficit shows that CAM induction is associated with the appearance of a molecular species of PEPC termed PEPC I. PEPC I is barely detectable in well-watered plants which are not performing CAM. The major form in these plants is termed PEPC II. In leaves from intact plants, there is a significant positive correlation between PEPC I activity and ΔH⁺ during a period of increasing water deficit. PEPC I exhibits day to night fluctuations in malate sensitivity, being less sensitive during the dark period. In contrast, PEPC II is more sensitive to inhibition by malate and has no day to night fluctuation in sensitivity. In detached leaves deprived of water, a small increase in PEPC I capacity is detected at the end of the first dark period (20 h after the start of treatment). The results suggest that PEPC I is required for attainment of maximum nocturnal malic acid synthesis. There is a significant correlation between leaf water status (relative water content), ΔH⁺, total PEPC and PEPC I activity suggesting that the internal water status of the plant may be a trigger for CAM induction. Abscisic acid applied to detached leaves does not cause nocturnal acidification.     

Evidence that the induction of crassulacean acid metabolism by water stress in Mesembryanthemum crystallinum (L.) involves root signalling

The aim of this study was to investigate whether the root system of Mesembryanthemum crystallinum (L.) plays a role in triggering the induction of crassulacean acid metabolism (CAM) during water stress. Depriving well-irrigated plants of water, by allowing the soil surrounding the roots to dry, caused increased daily losses in leaf relative water content (RVVC) and mesophyll cell turgor pressure. The RWC of the roots also declined. Subsequently plants exhibited physiological characteristics of CAM photosynthesis (i.e. diurnal fluctuations in leaf titratable acidity and nocturnal net CO₂ fixation). When the root system of plants was divided equally between two soil compartments and one half deprived of water, plants exhibited physiological characteristics of CAM without prior changes in leaf RWC content or mesophyll cell turgor pressure. Only the RWC of the water-stressed portion of the roots was reduced. These data suggest that in water-stressed plants daily changes in leaf water relations greater than those observed in well-irrigated plants, are not essential to trigger CAM expression. It is probable that a reduction in soil water availability can be perceived by the roots of M. crystallinum and that this information is conveyed to the leaves triggering the transition from C₃ to CAM photosynthesis.     

Oxygen isotope composition of CAM and C3 Clusia species: non-steady-state dynamics control leaf water 18O enrichment in succulent leaves

Leaf gas exchange and leaf water (18)O enrichment (Delta(18)O(L)) were measured in three Clusia species under field conditions during dry and wet seasons and in Miconia argentea during the dry season in the Republic of Panama. During the dry season, all three Clusia species used crassulacean acid metabolism (CAM); during the wet season Clusia pratensis operated in the C(3) mode, while Clusia uvitana and Clusia rosea used CAM. Large departures from isotopic steady state were observed in daytime Delta(18)O(L) of the Clusia species, especially during the dry season. In contrast, daytime Delta(18)O(L) was near isotopic steady state in the C(3) tree M. argentea. Across the full data set, non-steady-state predictions explained 49% of variation in observed Delta(18)O(L), whereas steady-state predictions explained only 14%. During the wet season, when Delta(18)O(L) could be compared with Clusia individuals operating in both C(3) and CAM modes, steady-state and non-steady-state models gave contrasting predictions with respect to interspecific variation in daytime Delta(18)O(L). The observed Delta(18)O(L) pattern matched that predicted for the non-steady state. The results provided a clear example of how non-steady-state control of leaf water (18)O dynamics can shift the slope of the relationship between transpiration rate and daytime Delta(18)O(L) from negative to positive.     

Daily fluctuations of titratable acidity, content of organic acids (malate and citrate) and soluble sugars of varieties and wild relatives of Ananas comosus L. growing under natural tropical conditions

The genus Ananas has its centre of origin in northern South America. In this area, several varieties of Ananas comosus are widely cultivated, and a number of wild species are found growing under variable conditions of light intensity, soil fertility and water availability. Here we report detailed daily courses of titratable acidity, and malate, citrate and free-sugars content of several cultivated varieties of A. comosus and of A. ananassoides, a closely related species growing on granitic rock-outcrops in southern Venezuela. Day-night oscillations of both malate and citrate were detected in plants growing under full sun, but malate was by far the most important organic anion associated with CAM performance in ail populations sampled. Fructose was the dominant compound in the neutral fraction, but only sucrose showed a consistent inverse relation with the cycle of titratable acidity. The diel oscillations of free sugars measured were not always enough to account for the amount of organic anions accumulated during the night. Plants cultivated under shady conditions always showed a lower night-time increase in titratable acidity and organic acids, and also smaller oscillations in the amount of free sugars than sun exposed plants. In all populations growing under full sun, osmolality increased during the night, but it was not always possible to explain these changes on the basis of variations in molar concentrations of organic acids and sugars. Besides, no diel variations in the cations K⁺, Ca²⁺ and Mg²⁺ were detected. K⁺ was always the dominant cation (K/Ca ratios ～ 19), while Mg²⁺ was always higher than Ca²⁺ (Mg/Ca ～ 2).