Modulation of photosynthesis and respiration in guard and mesophyll cell protoplasts by oxygen concentration

Stomatal movement is an energetic oxygen-requiring process. In the present study, the effect of oxygen concentration on mitochondrial respiratory activity and red-light-dependent photosynthetic oxygen evolution by Vicia faba and Brassica napus guard cell protoplasts was examined. Comparative measurements were made with mesophyll cell protoplasts isolated from the same species. At air saturated levels of dissolved oxygen in the protoplast suspension media, respiration rates by mesophyll protoplasts ranged from 6 to 10μmoles O₂ mg^?1 chl h^?1, while guard cell protoplasts respired at rates of 200–300 μmoles O₂ mg chl^?1 h^?1, depending on the species. Lowering the oxygen concentration below 50–60 mmol m^?3 resulted in a decrease in guard cell respiration rates, while rates by mesophyll cell protoplasts were reduced only at much lower concentrations of dissolved oxygen. Rates of photosynthesis in mesophyll cell protoplasts isolated from both species showed only a minor reduction in activity at low oxygen concentrations. In contrast, photosynthesis by guard cell protoplasts isolated from V. faba and B. napus decreased concomitantly with respiration. Oligomycin, an inhibitor of oxidative phos-phorylation, reduced photosynthesis in mesophyll cell protoplasts by 27–46% and in guard cell protoplasts by 51–58%. The reduction in both guard cell photosynthesis and respiration following exposure to low oxygen concentrations suggest close metabolic coupling between the two activities, possibly mediated by the availability of substrate for respiration associated with photosynthetic electron transport activity and subsequent export of redox equivalents.     

Acclimation of potato plants to polyethylene glycol-induced water deficit I. Photosynthesis and metabolism

The acclimation of photosynthesis and metabolism in response to water deficit is characterized using hydroponically grown potato plants (Solanum tuberosum cv. Désirée). Plants were subjected to a reduced water potential of the nutrient solution by adding 10% (w/v) PEG 6000. PEG-treated plants were retarded in growth. Leaves which had been fully developed before the PEG treatment and leaves grown during the PEG treatment showed different phenotypes and biochemical and physiological properties. Photosynthesis of all leaves decreased during the whole treatment. However, the decrease of photosynthesis in the two types of leaves had different causes indicated by differences in their metabolism. Leaves which were fully developed at the beginning of the PEG treatment began to wilt starting from the leaf rim. The apoplastic ABA content increased, coinciding with a decreased stomatal conductance. Increased energy charge of the cells indicated impaired chloroplastic metabolism, accompanied by a decrease of amounts of chloroplastic enzymes. The apoplastic and the symplastic ABA content were increased during water deficit and because ABA was concentrated in the cytosolic compartment it is suggested that ABA is involved in decreasing photosynthetic enzyme contents in old leaves. Young leaves, grown after the imposition of water deficit, were smaller than control leaves and had a curly surface. In young leaves apoplastic and cytosolic ABA contents were identical with control values. Carboxylation efficiency of photosynthesis was decreased, but the water use efficiency remained unchanged. Metabolic data of the photosynthetic pathways indicate a down-regulation of chloroplastic metabolism. It is concluded that in young leaves photosynthesis was non-stomatally limited. This limitation was not caused by ABA.     

Exaggerated root respiration accounts for growth retardation in a starchless mutant of Arabidopsis thaliana

The knock‐out mutation of plastidial phosphoglucomutase (pgm) causes a starchless phenotype in Arabidopsis thaliana, and results in a severe growth reduction of plants cultivated under diurnal conditions. It has been speculated that high soluble sugar levels accumulating during the light phase in leaf mesophyll might cause a reduction of photosynthetic activity or that shortage of reduced carbon during the night is the reason for the slow biomass gain of pgm. Separate simultaneous measurements of leaf net photosynthesis and root respiration demonstrate that photosynthetic activity per unit fresh weight is not reduced in pgm, whereas root respiration is strongly elevated. Comparison with a mutant defective in the dominating vacuolar invertase (AtβFruct4) revealed that high sucrose concentration in the cytosol, but not in the vacuole, of leaf cells is responsible for elevated assimilate transport to the root. Increased sugar supply to the root, as observed in pgm mutants, forces substantial respiratory losses. Because root respiration accounts for 80% of total plant respiration under long‐day conditions, this gives rise to retarded biomass formation. In contrast, reduced vacuolar invertase activity leads to reduced net photosynthesis in the shoot and lowered root respiration, and affords an increased root/shoot ratio. The results demonstrate that roots have very limited capacity for carbon storage but exert rigid control of supply for their maintenance metabolism.     

Effects of sulfur dioxide and ozone on intact leaves and isolated mesophyll cells of groundnut plants (Arachis hypogaea L.)

Difference between effects of sulfur dioxide (SO₂) and ozone (O₃) on groundnut plants (Arachis hypogaea L.) was studied by use of an exposure system of enzymatically-isolated mesophyll cells. SO₂ inhibited photosynthesis of intact groundnut leaves but induced no visible injury on leaves. SO₂ also inhibited photosynthesis of isolated mesophyll cells but did not kill the cells, suggesting that SO₂ inhibits photosynthesis by attacking rather specifically the photosynthetic apparatus in chloroplasts. O₃ inhibited photosynthesis of intact leaves and at the same time induced visible injury corresponding to the extent of photosynthesis inhibition. O₃ also inhibited photosynthesis of isolated mesophyll cells and killed the cells to the extent corresponding to photosynthesis inhibition, suggesting that O₃ inhibits photosynthesis not directly by attacking the photosynthetic apparatus but indirectly by killing cells. Since the response of intact leaves to each pollutant resembled that of isolated mesophyll cells, the difference between responses of intact leaves to both pollutants may considerably reflect that of mesophyll cells.     

Metabolic consequences of susceptibility and resistance (race-specific and broad-spectrum) in barley leaves challenged with powdery mildew

In a compatible interaction biotrophic fungi often lower the yield of their hosts by reducing photosynthesis and altering the fluxes of carbon within the infected leaf. In contrast, comparatively little is known about the metabolic consequences of activating resistance responses. In this study we investigated the hypothesis that the activation of both race-specific (Mla12) and broad-spectrum (mlo) resistance pathways in barley leaves infected with Blumeria graminis represents a cost to the plant in terms of carbon production and utilization. We have shown, using quantitative imaging of chlorophyll fluorescence, that during a susceptible interaction, photosynthesis was progressively reduced both in cells directly below fungal colonies and in adjacent cells when compared with uninoculated leaves. The lower rate of photosynthesis was associated with an increase in invertase activity, an accumulation of hexoses and a down-regulation of photosynthetic gene expression. During both Mla12- and mlo-mediated resistance, photosynthesis was also reduced, most severely inhibited in cells directly associated with attempted penetration of the fungus but also in surrounding cells. These cells displayed intense autofluorescence under ultraviolet illumination indicative of the accumulation of phenolic compounds and/or callose deposition. The depression in photosynthesis was not due only to cell death but also to an alteration in source-sink relations and carbon utilization. Apoplastic (cell wall-bound) invertase activity increased more rapidly and to a much greater extent than in infected susceptible leaves and was accompanied by an accumulation of hexoses that was localized to areas of the leaf actively exhibiting resistance responses. The accumulation of hexoses was accompanied by a down-regulation in the expression of Rubisco (rbcS) and chlorophyll a/b binding protein (cab) genes (although to a lesser extent than in a compatible interaction) and with an up-regulation in the expression of the pathogenesis-related protein 1 (PR-1). These results are consistent with a role for invertase in the generation of hexoses, which may supply energy for defence reactions and/or act as signals inducing defence gene expression.     

Expression of a yeast-derived invertase in the cell wall of tobacco and Arabidopsis plants leads to accumulation of carbohydrate and inhibition of photosynthesis and strongly influences growth and phenotype of transgenic tobacco plants.

Chimeric genes consisting of the coding sequence of the yeast invertase gene suc 2 and different N-terminal portions of the potato-derived vacuolar protein proteinase inhibitor II fused to the 35S CaMV promoter and the poly-A site of the octopine synthase gene were transferred into tobacco and Arabidopsis thaliana plants using Agrobacterium based systems. Regenerated transgenic plants display a 50- to 500-fold higher invertase activity compared to non-transformed control plants. This invertase is N-glycosylated and efficiently secreted from the plant cell leading to its apoplastic location. Whereas expression of the invertase does not lead to drastic changes in transgenic Arabidopsis thaliana plants, transgenic tobacco plants show dramatic changes with respect to development and phenotype. Expression of the invertase leads to stunted growth due to reduction of internodal distances, to development of bleached and/or necrotic regions in older leaves and to suppressed root formation. In mature leaves, high levels of soluble sugars and starch accumulate. These carbohydrates do not show a diurnal turnover. The accumulation of carbohydrate is accompanied by an inhibition of photosynthesis, and in tobacco, by an increase in the rate of respiration. Measurements in bleached versus green areas of the same leaf show that the bleached section contains high levels of carbohydrates and has lower photosynthesis and higher respiration than green sections. It is concluded that expression of invertase in the cell wall interrupts export and leads to an accumulation of carbohydrates and inhibition of photosynthesis.     

Effect of local irradiance on CO(2) transfer conductance of mesophyll in walnut

The acclimation responses of walnut leaf photosynthesis to the irradiance microclimate were investigated by characterizing the photosynthetic properties of the leaves sampled on young trees (Juglans nigraxregia) grown in simulated sun and shade environments, and within a mature walnut tree crown (Juglans regia) in the field. In the young trees, the CO(2) compensation point in the absence of mitochondrial respiration (Gamma*), which probes the CO(2) versus O(2) specificity of Rubisco, was not significantly different in sun and shade leaves. The maximal net assimilation rates and stomatal and mesophyll conductances to CO(2) transfer were markedly lower in shade than in sun leaves. Dark respiration rates were also lower in shade leaves. However, the percentage inhibition of respiration by light during photosynthesis was similar in both sun and shade leaves. The extent of the changes in photosynthetic capacity and mesophyll conductance between sun and shade leaves under simulated conditions was similar to that observed between sun and shade leaves collected within the mature tree crown. Moreover, mesophyll conductance was strongly correlated with maximal net assimilation and the relationships were not significantly different between the two experiments, despite marked differences in leaf anatomy. These results suggest that photosynthetic capacity is a valuable parameter for modelling within-canopies variations of mesophyll conductance due to leaf acclimation to light.     

Comparative studies of compatible and incompatible pepper–<Emphasis Type="Italic">Tobamovirus</Emphasis> interactions and the evaluation of effects of 24-epibrassinolide

The aim of study was to gain a deeper knowledge about local and systemic changes in photosynthetic processes and sugar production of pepper infected by Obuda pepper virus (ObPV) and Pepper mild mottle virus (PMMoV). PSII efficiency, reflectance, and gas exchange were measured 48 and/or 72 h after inoculation (hpi). Sugar accumulation was checked 72 hpi and 20 d after inoculation (as a systemic response). Inoculation of leaves with ObPV led to appearance of hypersensitive necrotic lesions (incompatible interaction), while PMMoV caused no visible symptoms (compatible interaction). ObPV (but not PMMoV) lowered F_v/F_m (from 0.827 to 0.148 at 72 hpi). Net photosynthesis decreased in ObPV-infected leaves. In ObPV-inoculated leaves, the accumulation of glucose, fructose, and glucose-6-phosphate was accompanied with lowered sucrose, maltoheptose, nystose, and trehalose contents. PMMoV inoculation increased the contents of glucose, maltose, and raffinose in the inoculated leaves, while glucose-6-phosphate accummulated in upper leaves.     

Systemic changes in photosynthesis and reactive oxygen species homeostasis in shoots of Arabidopsis thaliana infected with the beet cyst nematode Heterodera schachtii

Photosynthetic efficiency and redox homeostasis are important for plant physiological processes during regular development as well as defence responses. The second‐stage juveniles of Heterodera schachtii induce syncytial feeding sites in host roots. To ascertain whether the development of syncytia alters photosynthesis and the metabolism of reactive oxygen species (ROS), chlorophyll a fluorescence measurements and antioxidant responses were studied in Arabidopsis thaliana shoots on the day of inoculation and at 3, 7 and 15 days post‐inoculation (dpi). Nematode parasitism caused an accumulation of superoxide and hydrogen peroxide molecules in the shoots of infected plants at 3 dpi, probably as a result of the observed down‐regulation of antioxidant enzymes. These changes were accompanied by an increase in RNA and lipid oxidation markers. The activities of antioxidant enzymes were found to be enhanced on infection at 7 and 15 dpi, and the content of anthocyanins was elevated from 3 dpi. The fluorescence parameter R_fd, defining plant vitality and the photosynthetic capacity of leaves, decreased by 11% only at 7 dpi, and non‐photochemical quenching (NPQ), indicating the effectiveness of photoprotection mechanisms, was about 16% lower at 3 and 7 dpi. As a result of infection, the ultrastructure of chloroplasts was changed (large starch grains and plastoglobules), and more numerous and larger peroxisomes were observed in the mesophyll cells of leaves. We postulate that the joint action of antioxidant enzymes/molecules and photochemical mechanisms leading to the maintenance of photosynthetic efficiency promotes the fine‐tuning of the infected plants to oxidative stress induced by parasitic cyst nematodes.     

Effects of light and temperature on leaf anatomy and photosynthesis in Fragaria vesca

Summary Fragaria vesca, the woodland strawberry, was grown under a series of controlled environments including variations in light intensity, average temperatures, and temperature amplitude around a constant mean. Observations on CO₂ exchange capacities, leaf anatomy, and cell ultrastructure were made for each treatment to determine relationships between these variables. With increasing light intensity, leaf thickness, leaf density, and mesophyll cell surface area and volume per leaf surface area increased. Net photosynthesis (NPS) per leaf weight decreased with increasing light pretreatment while NPS per area increased from low to medium intensity, then decreased at the highest intensity. Depression of photosynthesis at the highest light pretreatment may have been due to massive starch accumulation in the chloroplasts associated with the sodium vapor lamps used. Correlation of all anatomical variables was highly significant with dark respiration and NPS per dry weight but insignificant for NPS per leaf area. In the variable temperature treatments, photosynthetic acclimation occurred with a shift in optimum temperature for NPS in the direction of prevailing growth temperature. Absolute rates were highest at moderate pretreatment temperatures and were reduced by extreme growth temperatures. Thick leaves with low density mesophyll became thinner and more dense with increasing growth temperature corresponding to an increase in maximum net photosynthetic rates. Leaves became thicker and more dense at the highest temperatures, but with an increase in cell damage and indications of changes in metabolic pathways. Highest correlations for gas exchange rates were with specific leaf weight (weight per area). Correlation with other anatomical variables were scattered or insignificant. It was concluded that adaptation to a range of environmental conditions cannot be consistently attributed to changes in mesophyll cell volume or surface area.     

The role of mesophyll conductance in the economics of nitrogen and water use in photosynthesis

A recent resurgence of interest in formal optimisation theory has begun to improve our understanding of how variations in stomatal conductance and photosynthetic capacity control the response of whole plant photosynthesis and growth to the environment. However, mesophyll conductance exhibits similar variation and has similar impact on photosynthesis as stomatal conductance; yet, the role of mesophyll conductance in the economics of photosynthetic resource use has not been thoroughly explored. In this article, we first briefly summarise the knowledge of how mesophyll conductance varies in relation to environmental factors that also affect stomatal conductance and photosynthetic capacity, and then we use a simple analytical approach to begin to explore how these important controls on photosynthesis should mutually co-vary in a plant canopy in the optimum. Our analysis predicts that when either stomatal or mesophyll conductance is limited by fundamental biophysical constraints in some areas of a canopy, e.g. reduced stomatal conductance in upper canopy leaves due to reduced water potential, the other of the two conductances should increase in those leaves, while photosynthetic capacity should decrease. Our analysis also predicts that if mesophyll conductance depends on nitrogen investment in one or more proteins, then nitrogen investment should shift away from Rubisco and towards mesophyll conductance if hydraulic or other constraints cause chloroplastic CO₂ concentration to decline. Thorough exploration of these issues awaits better knowledge of whether and how mesophyll conductance is itself limited by nitrogen investment, and about how these determinants of photosynthetic CO₂ supply and demand co-vary among leaves in real plant canopies.     

Photosynthetic limitations in olive cultivars with different sensitivity to salt stress

Olive (Olea europea L) is one of the most valuable and widespread fruit trees in the Mediterranean area. To breed olive for resistance to salinity, an environmental constraint typical of the Mediterranean, is an important goal. The photosynthetic limitations associated with salt stress caused by irrigation with saline (200 mm ) water were assessed with simultaneous gas‐exchange and fluorescence field measurements in six olive cultivars. Cultivars were found to possess inherently different photosynthesis when non‐stressed. When exposed to salt stress, cultivars with inherently high photosynthesis showed the highest photosynthetic reductions. There was no relationship between salt accumulation and photosynthesis reduction in either young or old leaves. Thus photosynthetic sensitivity to salt did not depend on salt exclusion or compartmentalization in the old leaves of the olive cultivars investigated. Salt reduced the photochemical efficiency, but this reduction was also not associated with photosynthesis reduction. Salt caused a reduction of stomatal and mesophyll conductance, especially in cultivars with inherently high photosynthesis. Mesophyll conductance was generally strongly associated with photosynthesis, but not in salt‐stressed leaves with a mesophyll conductance higher than 50 mmol m^?2 s^?1. The combined reduction of stomatal and mesophyll conductances in salt‐stressed leaves increased the CO₂ draw‐down between ambient air and the chloroplasts. The CO₂ draw‐down was strongly associated with photosynthesis reduction of salt‐stressed leaves but also with the variable photosynthesis of controls. The relationship between photosynthesis and CO₂ draw‐down remained unchanged in most of the cultivars, suggesting no or small changes in Rubisco activity of salt‐stressed leaves. The present results indicate that the low chloroplast CO₂ concentration set by both low stomatal and mesophyll conductances were the main limitations of photosynthesis in salt‐stressed olive as well as in cultivars with inherently low photosynthesis. It is consequently suggested that, independently of the apparent sensitivity of photosynthesis to salt, this effect may be relieved if conductances to CO₂ diffusion are restored.     

Cell wall-bound invertase limits sucrose export and is involved in symptom development and inhibition of photosynthesis during compatible interaction between tomato and Xanthomonas campestris pv vesicatoria

Cell wall-bound invertase (cw-Inv) plays an important role in carbohydrate partitioning and regulation of sink-source interaction. There is increasing evidence that pathogens interfere with sink-source interaction, and induction of cw-Inv activity has frequently been shown in response to pathogen infection. To investigate the role of cw-Inv, transgenic tomato (Solanum lycopersicum) plants silenced for the major leaf cw-Inv isoforms were generated and analyzed during normal growth and during the compatible interaction with Xanthomonas campestris pv vesicatoria. Under normal growth conditions, activities of sucrolytic enzymes as well as photosynthesis and respiration were unaltered in the transgenic plants compared with wild-type plants. However, starch levels of source leaves were strongly reduced, which was most likely caused by an enhanced sucrose exudation rate. Following X. campestris pv vesicatoria infection, cw-Inv-silenced plants showed an increased sucrose to hexose ratio in the apoplast of leaves. Symptom development, inhibition of photosynthesis, and expression of photosynthetic genes were clearly delayed in transgenic plants compared with wild-type plants. In addition, induction of senescence-associated and pathogenesis-related genes observed in infected wild-type plants was abolished in cw-Inv-silenced tomato lines. These changes were not associated with decreased bacterial growth. In conclusion, cw-Inv restricts carbon export from source leaves and regulates the sucrose to hexose ratio in the apoplast. Furthermore, an increased apoplastic hexose to sucrose ratio can be linked to inhibition of photosynthesis and induction of pathogenesis-related gene expression but does not significantly influence bacterial growth. Indirectly, bacteria may benefit from low invertase activity, since the longevity of host cells is raised and basal defense might be dampened.     

Stomatal aperture,photosythesis and water fluxes in mesophyll cells as affected by the abscission of leaves. Simultaneous measurements of gas exchange,light scattering and chlorphyll fluorescence

Carbon dioxide exchange, transpiration, chlorophyll fluorescence and light scattering of leaves of Lycopersicom esculentum, Helianthus annuus and Arbutus unedo were measured simultaneously before and after abscission of leaves. Scattering of a weak green measuring beam was used to monitor water fluxes across the thylakoid membranes of the mesophyll. When leaves were cut under water, stomata initially closed partially and then occasionally exhibited distinct regulatory oscillations. As stomata closed, light scattering decreased indicating water influx into the mesophyll. Stomatal oscillations were accompanied, with small but noticeable phase shifts, by oscillations of water fluxes at the thylakoid level. These fluxes could be distinguished from the water fluxes accompanying light-dependent ion pumping across the thylakoids by the concomitant chlorophyll fluorescence signals. The latter record energy-dependent ion fluxes in addition to redox changes of the electron-transport chain. As stomata closed partially after cutting a leaf under water, photosynthesis decreased. In Arbutus unedo and Helianthus annuus leaves, transient stomatal closure was insufficient to account for transient inhibition of photosynthesis which appeared to be brought about by transfer of an inhibitory solute through the petiole into the mesophyll. This solute also stimulated respiration in the dark. When leaves were cut in air, stomata opened transiently (Iwanoff effect) before wilting enforced closure. Photosynthesis followed the stomatal responses, increasing during opening and decreasing during closure.Dedicated to Professor H. Ullrich on the occasion of his 85th birthday     

Antioxidant defences of the apoplast

Summary The apoplast of barley and oat leaves contained superoxide dismutase (SOD), catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase activities. The activities of these enzymes in the apoplastic extracts were greatly modified 24 h after inoculation with the biotrophic fungal pathogenBlumeria graminis. The quantum efficiency of photosystem II, which is related to photosynthetic electron transport flux, was comparable in inoculated and healthy leaves during this period. Apoplastic soluble acid invertase activity was also modified in inoculated leaves. Inoculation-dependent increases in apoplastic SOD activity were observed in all lines. Major bands of SOD activity, observed in apoplastic protein extracts by activity staining of gels following isoelectric focusing, were similar to those observed in whole leaves but two additional minor bands were found in the apoplastic fraction. The apoplastic extracts contained substantial amounts of dehydroascorbate (DHA) but little or no glutathione (GSH). Biotic stress decreased apoplastic ascorbate and DHA but increased apoplastic GSH in resistant lines. The antioxidant cycle enzymes may function to remove apoplastic H₂O₂ with ascorbate and GSH derived from the cytoplasm. DHA and oxidized glutathione may be reduced in the apoplast or returned to the cytosol for rereduction.Abbreviations AA reduced ascorbate - APX ascorbate peroxidase - DHA dehydroascorbate (oxidised ascorbate) - DHAR dehydroascorbate reductase - G6PDH glucose-6-phosphate dehydrogenase - GSH reduced glutathione - GSSG glutathione disulphide - GR glutathione reductase - MDHA monodehydroascorbate - MDHAR monodehydroascorbate reductase - SOD superoxide dismutase     

Seasonal changes in photosynthetic characteristics of Anemone raddeana,a spring-active geophyte,in the temperate region of Japan

Summary Seasonal changes in the photosynthetic characteristics of intact involucral leaves of Anemone raddeana were investigated under laboratory conditions. Net photosynthesis and constant water vapor pressure deficit showed almost the same seasonal trend. They increased rapidly from mid-April immediately after unfolding of the leaves and reached the maximum in late-April, before the maximum expansion of the leaves. They retained the maximum values until early-May and then decreased toward late-May with a progress of leaf senescence. The calculated values of intercellular CO₂ concentration and relative stomatal limitation of photosynthesis showed no significant change throughout the season. The carboxylation efficiency as assessed by the initial slope of Ci-photosynthesis curve and the net photosynthesis under a high Ci regime varied seasonally in parallel with the change of the light-saturated photosynthesis. The results indicate that the seasonal changes in light-saturated net photosynthesis are not due to a change of stomatal conductance, but to a change in the photosynthetic capacity of mesophyll. Nevertheless, leaf conductance changed concomitantly with photosynthetic capacity, indicating that the seasonal change in stomatal conductance is modulated by the mesophyll photosynthetic capacity such that the intercellular CO₂ concentrations is maintained constant. The shape of light-photosynthesis curve was similar to that of sun-leaf type. The quantum yield also changed simultaneously with the photosynthetic capacity throughout the season.Contribution No. 2965 from the Institute of Low Temperature Science     

Role of the Apoplast in the Control of Assimilate Transport,Photosynthesis, and Plant Productivity

A concept is suggested, which supposes that assimilates are transferred within the plant downward through phloem sieve tubes and, after entering the stem apoplast, are carried up with the ascending flow of transpiration water. After entering the apoplast of fully expanded leaves, these solutes are reexported through the phloem. Thus, a common pool of assimilates with uniform concentration is formed in the plant apoplast. According to this concept, the mechanism of assimilate demand represents a response of photosynthetic apparatus to changes in the apoplastic level of metabolites consumed by sink organs. The ratios of labeled photoassimilates differ between the apoplast and mesophyll cells. Most of the apoplastic labeled carbon is contained in sucrose, less in amino acids, and even less in hexoses. The ¹⁴C-labeling of amino acids increases and the sucrose/hexose labeling ratio decreased under conditions of enhanced nitrate supply. The well-known effect of relative inhibition of assimilate export from leaves under conditions of enhanced nitrogen supply is explained by an enhanced hydrolysis of apoplast-derived sucrose due to the increase in invertase activity, rather than by diversion of primary photosynthetic products from sucrose synthesis to other pathways required for activated growth processes in leaves. This notion is based on observations that the sucrose/hexose ratio is reduced to a greater extent in the apoplast than in the symplast. The last assumption was supported by data obtained after artificial changes in the apoplastic pH. In these experiments intact plants were placed in the atmosphere of NH₃ or HCl vapors, which induced opposite changes in relative content of labeled assimilates in the apoplast and in the photosynthetic rate.     

The Decreased Cold-Resistance of Chilling-Sensitive Plants Is Related to Suppressed CO2 Assimilation in Leaves and Sugar Accumulation in Roots

Tomato (Lycopersicon esculentum L., cv. Sibirskii skorospelyi) and cucumber (Cucumis sativus L., cv. Konkurent) plants were grown in a soil culture in a greenhouse at an average daily temperature of 20°C and ambient illumination until the development of five and eight true leaves, respectively. During the subsequent three days, some plants were kept in a climatic chamber at 6°C in the light, whereas other plants remained in a greenhouse (control). The cold-resistance of cucumber leaves and roots, as assayed from the electrolyte leakage, was reduced after cold exposure stronger than cold-resistance of tomato organs. The ratio photosynthesis/dark respiration was lower in cucumber than in tomato leaves at all measurement temperatures. The concentrations of sugars (sucrose + glucose + fructose) increased in chilled tomato roots but decreased in cucumber roots. Cold exposure changed the activities of various invertase forms (soluble and insoluble acidic and alkaline invertases). The total invertase activity and the ratio of mono- to disaccharides increased. The lower cucumber cold-resistance is related to the higher sensitivity of its photosynthetic apparatus to chilling and, as a consequence, insufficient root supply with sugars.     

Photosystem II efficiency of the palisade and spongy mesophyll in <Emphasis Type="Italic">Quercus coccifera</Emphasis> using adaxial/abaxial illumination and excitation light sources with wavelengths varying in penetration into the leaf tissue

The existence of major vertical gradients within the leaf is often overlooked in studies of photosynthesis. These gradients, which involve light heterogeneity, cell composition, and CO₂ concentration across the mesophyll, can generate differences in the maximum potential PSII efficiency (F _V/F _M or F _V/F _P) of the different cell layers. Evidence is presented for a step gradient of F _V/F _P ratios across the mesophyll, from the adaxial (palisade parenchyma, optimal efficiencies) to the abaxial (spongy parenchyma, sub-optimal efficiencies) side of Quercus coccifera leaves. For this purpose, light sources with different wavelengths that penetrate more or less deep within the leaf were employed, and measurements from the adaxial and abaxial sides were performed. To our knowledge, this is the first report where a low photosynthetic performance in the abaxial side of leaves is accompanied by impaired F _V/F _P ratios. This low photosynthetic efficiency of the abaxial side could be related to the occurrence of bundle sheath extensions, which facilitates the penetration of high light intensities deep within the mesophyll. Also, leaf morphology (twisted in shape) and orientation (with a marked angle from the horizontal plane) imply direct sunlight illumination of the abaxial side. The existence of cell layers within leaves with different photosynthetic efficiencies makes appropriate the evaluation of how light penetrates within the mesophyll when using Chl fluorescence or gas exchange techniques that use different wavelengths for excitation and/or for driving photosynthesis.     

Absence of turnover and futile cycling of sucrose in leaves of<Emphasis Type="Italic"> Lolium temulentum</Emphasis> L.: implications for metabolic compartmentation

To study the interdependence of sucrose accumulation and its hydrolyzing enzyme, soluble acid invertase (AI; EC 3.2.1.26), in fructan-accumulating temperate grasses and cereals, experiments were performed in which sucrose synthesis was abolished in leaves of Lolium temulentum by four independent inhibitory factors, each having a distinct mechanism of action. Trials in the light with mannose or vanadate and in the dark with anoxia or cyanide showed that previously accumulated sucrose was stable in the tissue over a 5- to 6-h period. Conversely, putatively vacuolar AI activity in tissue homogenates was sufficient to completely convert endogenous sucrose to monosaccharide within the same period. Continuous invertase-mediated breakdown of sucrose was thus not a feature of this tissue. It is concluded that AI and sucrose were not in metabolic contact in vivo, implying differential compartmentation. In darkness, in uninhibited leaves, sucrose concentrations fell linearly with respect to time at a rate of –0.6 mg g^–1 FW h^–1, over a 5- to 6-h period. This value is equivalent to rates of dark respiration measured by gas exchange. Dark-utilisation of sucrose was not accompanied by monosaccharide accumulation in the tissue. The rate of sucrose loss was 3-fold lower than rates of extractable AI activity. Hence, if AI was involved in dark-utilisation, then this implies at least a partial differential localisation of enzyme and substrate. However, the dark-consumption of sucrose was completely abolished by anoxia and by cyanide. It follows that dark-mobilisation (unlike invertase hydrolysis per se) was respiration-dependent and did not result from a simple co-localisation of sucrose and invertase. Taken together, the results show that sucrose and invertase do not share the same metabolic compartment in grass leaves. It is possible that invertase has no role in the mobilisation of stored sucrose in leaves of the fructan-accumulating grasses.Abbreviations AI Acid invertase - PAR Photosynthetically active radiation - TLC Thin-layer chromatography