Carbon dioxide concentration and nitrogen input affect the C and N storage pools in Amanita muscaria-Picea abies mycorrhizae.

We studied the influence of elevated atmospheric CO2 concentration ([CO2]) on the vacuolar storage pool of nitrogen-containing compounds and on the glycogen pool in the hyphal sheath of Amanita muscaria (L. ex Fr.) Hooker-Picea abies L. Karst. mycorrhizae grown with two concentrations of ammonium in the substrate. Mycorrhizal seedlings were grown in petri dishes on agar containing 5.3 or 53 mg N l(-1) and exposed to 350 or 700 microl CO2 l(-1) for 5 or 7 weeks, respectively. Numbers and area of nitrogen-containing bodies in the vacuoles of the mycorrhizal fungus were determined by light microscopy linked to an image analysis system. The relative concentration of nitrogen in the vacuolar bodies was measured by electron energy loss spectroscopy (EELS). Glycogen stored in the cytosol was determined at the ultrastructural level by image analysis after staining the sections (PATAg test). Shoot dry weight, net photosynthesis and relative amounts of N in vacuolar bodies were greater at the higher N and CO2 concentrations. The numbers and areas of vacuolar N-containing bodies were significantly greater at the higher N concentration only at ambient [CO2]. In the same treatment the percentage of hyphae containing glycogen declined to nearly zero. We conclude that, in the high N/low [CO2] treatment, the mycorrhizal fungus had an insufficient carbohydrate supply, partly because of increased amino acid synthesis by the non-mycorrhizal rootlets. When [CO2] was increased, the equilibrium between storage of glycogen and N-containing compounds was reestablished.

Effect of fertilization on ozone-induced changes in the metabolism of birch (Betula pendula) leaves.

Cloned cuttings of Betula pendula Roth were grown in field fumigation chambers at Birmensdorf throughout one growing season in filtered air with either < 3 (control) or 90/40 nl l-1 O3 (day/night; ozone generated from pure oxygen). Each ozone regime was split into high and low soil nutrient regimes by watering plants with either a 0.05 % or a 0.005% solution of a fertilizer which contained macronutrients and micronutrients. Fertilization had a strong effect on plant growth, enzyme activities and the expression of ozone-induced effects at the biochemical level. The activities of PEPC and Rubisco were enhanced about threefold in the plants with high fertilization (HF). Significant effects of ozone were in most cases found only in the older leaves of the plants with low fertilization (LF), There, sucrose, glucose and fructose levels were enhanced. In both fertilization treatments, the number of starch granules along the minor veins was increased. These ozone effects point to a decreased or inhibited phloem loading. The increased PEPC activity and the enhanced malate levels in the ozone-exposed plants might be the result of a redirection of carbon flow from sucrose synthesis and translocation towards anapleurotic processes, which can feed detoxification and repair of ozone injury as indicated by enhanced respiration. These findings agree well with the observed effects of ozone in lowering the root: shoot biomass ratio. Although there was a marked reduction in the O3 /LF plants, O3 /HF plants showed no significant response. Inositol was decreased under ozone exposure in both fertilizer treatments, contrasting with the pattern for carbohydrates. These results demonstrate the role of fertilization as an important modifier of ozone-induced effects at the plant biochemical level. Well fertilized plants appear to cope better with the impact of ozone on metabolism.

Coarse and Fine Control and Annual Changes of Sucrose-Phosphate Synthase in Norway Spruce Needles.

Annual changes of activity of sucrose-phosphate synthase (SPS) from spruce (Picea abies [L.] Karst.) needles were studied with respect to three regulatory levels: metabolic fine control, covalent modification (phosphorylation), and protein amount. Glucose-6-phosphate served as an allosteric activator of spruce SPS by shifting the Michaelis constant for the substrate fructose-6-phosphate from 4.2 to 0.59 mM, whereas inorganic phosphate competitively inhibited this activation. The affinity for the other substrate, UDP-glucose, was unaffected. Incubation of the crude extract with ATP resulted in a time- and concentration-dependent decrease of the maximal velocity of SPS. This inactivation was sensitive to staurosporine, a potent protein kinase inhibitor, indicating the participation of a protein kinase. Probing SPS protein with heterologous antibodies showed that the subunit of spruce SPS is an approximately 139-kD protein and that changes in the extractable activity during the course of a year were correlated with the amount of SPS protein. High SPS activities in winter were paralleled by increased levels of the activator glucose-6-phosphate and the substrate fructose-6-phosphate, indicating a high capacity for sucrose synthesis that may be necessary to maintain photosynthetic CO2 fixation in cold-hardened spruce needles.

Energy and redox status, and carbon allocation in one- to three-year-old spruce needles.

Metabolite pools were determined in lyophilized homogenates of one- to three-year-old needles from 13-year-old spruce trees. These were exposed in open-chambers for five consecutive years to SO(2) (30 microg m(-3)) and O(3) (50 to 180 microg m(-3)), singly and in combination. The samples were taken shortly before bud break (mid-March). The energy (ATP/ADP) and redox status (NAD(P)H/NAD(P)) increased with needle age. This increase was most pronounced in needles exposed to both pollutant gases. To evaluate carbohydrate metabolism, levels of starch, sugars (sucrose, fructose, glucose), sugar phosphates (dihydroxyacetone phosphate, glyceraldehyde 3-phosphate (triose phosphates, TP), fructose 6-phosphate, fructose 2,6-bisphosphate: F2,6BP) and 3-phosphoglyceric acid (PGA) were measured. The concentrations of starch, glucose, fructose and TP on a dry weight basis increased with needle age, while those of sucrose and F2,6BP decreased. Interestingly, the total amount of soluble sugars remained constant in needles from control chambers and large fluctuations mainly occurred after a single application of the pollutants. Needles from fumigated trees generally showed somewhat lower starch and sugar, and fructose 6-phosphate levels. The content of TP increased with needle age while that of sucrose and F2,6BP decreased. This inverse behavior was even more prominent in the presence of both pollutants and is discussed in the context of recent knowledge on the regulation of carbon partitioning. There was a significant increase in the ratio of TP/PGA in samples from the combined exposure treatments. This also indicates an increase in the redox charge and corroborates the data on both the redox status of the pyridine nucleotide system and the significantly increased ATP/ADP ratio. The results are indicative of a switch in metabolism, from anabolic to predominantly catabolic reactions, which is most pronounced in the samples fumigated with both O(3) and SO(2). The data are compared to those obtained from naturally exposed trees in the southern Black Forest (Kälbelescheuer/Haldenhof, near Freiburg, Germany).