Günter Döhler

Impact of UV-B Radiation on Photosynthetic Assimilation of 14C-Bicarbonate and Inorganic 15N-Compounds by Cyanobacteria

The cyanobacteria Anabaena cylindrica and Synechococcus leopoliensis (= Anacystis nidulans) were grown at different levels of UV-B radiation (439. 717, 1230 and 1405 J m -2d-1 weighted according Caldwell, 1971) for 2 days. Dry weight was hardly affected but phycocyanin content of both species decreased linearly to the level of UV-B radiation. Contents of protein, carotenoids and chlorophyll a were reduced only after exposure to high doses (1230 J m-2d-1) of UV-B radiation. Photosynthetic 14CO2 fixation of Anabaena cells was reduced linearly with increasing UV-B dose whereas no effect could be observed in Synechococcus. A depression of photosynthetic 15N-nitrate uptake was found after UV-B stress in both species. UV-B irradiance caused an increase of 15N-incorporation into glutamine, but no effect was noted for incorporation into alanine or aspartic acid. An increase of 15N-excess in glutamic acid linear with the UV-B dose was observed in Synechococcus, only. Patterns of 14C-labelled photosynthetic products were either less affected by UV-B radiation (Anabaena) or an enhancement of 14C-label in total amino acids was detected (Synechococcus). The amount of total free amino acids increased parallel to the level of UV-B radiation. Only, the high dose of UV-B (1405 J m-2d-1, weighted) results in a decrease of the glutamine pool. Our results indicate an inhibition of glutamate synthase by UV-B irradiation in Anabaena, only. Results were discussed with reference to the damage of the photosynthetic apparatus.

Impact of UV-B Radiation on the Lipid and Fatty Acid Composition of Synchronized Ditylum brightwellii (West) Grunow

Abstract The marine diatom Ditylum brightwellii (West) Grunow isolated from the Baltic Sea could be synchronized by a light/dark rhythm of 6.5:17.5 h (white light intensity 8 W m-2) at 18 °C and 0.035 vol.% CO2. Content of protein, DNA and RNA increased linearly up to the end of the cell cycle. Pigments (chlorophyll a, chlorophyll c1 + c2, carotenoids) and galactolipids were synthesized in the light period only. A lag phase of 2 h was observed in the biosynthesis of sulphoquinovosyl diacylglycerol and phosphatidylglycerol. Formation of phosphatidylglycerol and phosphatidylcholin continued in the dark period (30% and 28%, respectively). The pattern of major fatty acids (C14:0, C16:1, C16:0, C18:1 and C20:5) varied during the cell cycle of Ditylum. Biosynthesis of acyl lipids was reduced in dependence on the UV-B dose. The most sensitive lipid was digalactosyl diacylglycerol (total inhibition at 585 J m-2), whereas phosphatidylcholin was less affected (20% reduction). UV-B radiation during the dark period had no effect on the lipid and pigment content. Strongest inhibitory effect of UV-B on cell division, synthesis of protein, pigments, sulphoquinovosyl diacylglycerol and phosphatidylglycerol was found after UV-B radiation at the beginning of the cell cycle (0.-2. h). An exposure time at the end of the light period (4.-6. h) led to a marked damage on the synthesis of monogalactosyl diacylglycerol and phosphatidylglycerol. These findings indicate a stage-dependent response of Ditylum to UV-B irradiance. The impact of UV-B resulted in an increase of unsaturated long chained fatty acids (C18, C20) and in a diminution of short chained fatty acids (C14, C16). Content of ATP was not affected by UV-B radiation under the used conditions. The inhibitory effect of UV-B on synthesis of DNA, RNA, protein and acyl lipids was mainly reversible. Results were discussed with reference to UV-B damage on the enzymes involved in the biosynthesis of acyl lipids and by a reduction of available metabolites.

Einfluß der Kulturbedingungen auf die photosynthetischen Carboxylierungsreaktionen von Chlorella vulgaris

Summary CO 2 exchange, 14 CO 2 fixation and 14 C-incorporation in photosynthetic products of Chlorrella vulgaris (strain 211–11f) were studied during the photosynthetic induction period at temperatures of + 10° C and + 35° C. The algae were grown in low (0.035 vol. %) and high (3.5 vol. %) CO 2 concentrations and at different temperatures (+ 15°, + 25° and + 35° C). Transient changes in CO 2 uptake, measured with an infrared gas analyzer, are very dependent on the CO 2 contents and temperatures during both the growth and the measurements. The maximum in CO 2 uptake at the beginning of the photosynthetic period decreases, when the algae were grown in high CO 2 concentrations. The autoradiographic studies of the kinetics of the appearance of labelled products at + 10° C and + 35° C showed that in Chlorella cells grown in both CO 2 concentrations and at growth temperatures of + 15° C and + 35° C the Calvin cycle is the main carboxylation reaction. At the beginning of the illumination period in Chlorella cells grown at + 25° C 14 CO 2 was mainly incorporated into malate, aspartate and 3-phosphoglycerate at a temperature of +10° C. High CO 2 concentrations during the growth phase of the algae had only a small effect on the percentage distribution of relabelled products. Furthermore, we estimated the enzyme activities of phosphoenolpyruvat carboxylase, malate dehydrogenase, malic enzyme and ribulose-1,5-diphosphate carboxylase in in cell-free extracts of Chlorella vulgaris grown under different conditions. A very active PEP-carboxylase could be found in extracts of Chlorella grown at + 25° C in air and 3.5 vol. % CO 2 . We observed a higher activity of RuDP-carboxylase in air-grown Chlorella cells as compared to CO 2 -grown cells. The results showed that the temperature during the growth of algae affects the carbon dioxide fixation and the enzyme activities more than the CO 2 concentration.

Effect of UV-B radiation on biomass production, pigmentation and protein content of marine diatoms

Several species of marine diatoms were grown at + 18 °C and + 22 °C under normal air conditions (0.035 vol.% C02) at a light/dark alteration of 14: 8̄ h. Intensity of white light was 1 mW (~ 5000 lux). An artifical nutrient solution of 35%o salinity was used. Algae - harvested during exponential growth - were exposed to different intensities of UV-B radiation (439, 717 and 1230 J · m-2 · m-1) for 2 days. UV-B radiation depressed the growth of all tested marine diatoms. Low levels of UV-B resulted in a slight increase of the biomass production (dry weight) compared to not UV-B treated cells. Enhanced UV-B doses caused a diminution of the primary productivity in all species. Algae exposed to UV-B stress showed a marked decrease in the protein and pigment content (chlorophyll a, chlorophyll c1 + c2 and carotenoids). In + 22°C grown cells of Lauderia annulata and Thalassiosira rotula were more sensitive to UV-B radiation than those cultures grown at + 18 °C. Bellerochea yucatanensis cells grown at +22 °C were less affected after UV-B exposure than at +18°C grown algae. The UV-B sensibility and growth of the individual species varied in a mixture of several marine diatoms. Results were discussed with reference to the UV-B effect on metabolic processes.

Photosynthetic Assimilation of 15N-Ammonia and 15N-Nitrate in the Marine Diatoms Bellerochea yucatanensis (von Stosch) and Skeletonema costatum

Abstract 14C-and 15N-Assimilation, 15N-Labelled Amino Acids, M arine D iatom s The marine diatoms Bellerochea yucatanensis and Skeletonema costatum were grown at +20 °C in 0.03 vol.% CO2 with nitrate or ammonia. The 15N -am m onia and 15N -nitrate assim ila tion and 15N -incorporation into various amino acids were studied of both diatom s during exponential growth phase in dependence of different nitrogen conditions. In all experiments the 15N -am m onia uptake was lower than the 15N -nitrate assim ilation rate up to 20-40 min photo synthesis. N itrate lim itation -cells grown in nitrate followed by growth in nitrogen-free m edium for 24 h — caused a strong 15N-label into aspartate after adding 15NH 4C1 (1 m M). In cells grown in nitrate highest enrichment of 15N was found in glutamine. Results were discussed with reference to the operating of the GS/GOGAT system and glutam ic acid dehydrogenase pathway. Photosynthetic 14CO2 fixation experiments showed a very high labelling of aspartate which was interpreted with a phosphoenolpyruvate carboxylation catalysed by phosphoenolpyruvate carb-oxykinase.

Effect of Light on Lipid and Fatty Acid Composition of Cyanobacteria, Anacystis nidulans (Synechococcus)

Summary Pigment alterations could be obtained by exposing Anacystis cultured under low white light (0.6 X 103 µW/cm2) to high white light (3 X 104 µW/cm2) or high red light (2 X 104 µW/cm2). The amounts of total pigments and acyl lipids decreased from red light to high white light grown cells. Within the different cultures a relative constant proportion of lipid soluble pigments and acyl lipids could be observed. Regarding to the influence of culture conditions on the composition and percentage distribution of individual aryl lipids no effect on SQDG could be noted but high light irrespective of light quality leads to a slight -decrease in PG content. Galactolipids and the MGDG/DGDG-ratio proved to be dependent on the chosen illumination. High red light yielded in an increase in MGDG synthesis, while high white light dependent lipid decrease was lesser for DGDG than for all other lipids. In high white light grown cultures a glycolipid, not yet reported to occur in Anacystis nidulans or other cyanobacteria was detected. Concerning the fatty acid composition, this lipid shows the lowest degree of desaturation of all lipids investigated. The sugar moiety of this unknown lipid is still to be researched. Common for all cultures the galactolipids can be divided from SQDG and PG in the percentage distribution of their faty acids. C16 : 0, C16 : 1 and C18 : 1 as major components in all lipids show little to, no difference in percentage distribution between cyanobacteria grown in low white and high red light. High white light led to a relative higher amount of C16 : 0 and C18 : 1 in all acyl lipids, whereas C18 : 1 correspondingly decreased. As minor fatty acid components in lipids of all cultures C14 :0 C14 : 11 C17 :15 C18 : 0 and C20 : 0 were observed.

Photosynthetische carboxylierungsreaktionen synchron kultivierter Chlorella vulgaris

Summary CO 2 exchange, 14 CO 2 fixation and 14 C-labelled photosynthetic products were studied in synchronized cultures of Chlorella vulgaris (strain 211-11 f) during the photosynthetic induction period. The algae were grown in 2,0 vol.% CO 2 at 25° C and at a periodic light dark regime with 16 hours of light and 8 hours of darkness. Transient changes in CO 2 up-take,measured with an infrared gas analyzer, varied over the division cycle. A maximum in CO 2 uptake at the beginning of the photosynthetic period exists only in autosporemother cells. The autoradiographic studies of the kinetics of the appearance of labelled products at + 25° C showed that both the total CO 2 fixation and the labelling of the different intermediates varied with the age of the Chlorella cells. The Calvin cycle is the main carboxyla-tion reaction in autospores. However, the percentage distribution of the labelled photosynthetic products of autosporemother cells shows that the C 4 dicarboxylic acids are primary products of CO 2 -fixation in light. A light induced glycolate release could be observed in all investigated stages of the division cycle. The labelling of glycolate was in autospores higher than in autosporemother cells. The specific activities of phosphoenolpyruvate carboxylase, malate dehydrogenase, malic enzyme, ribulose-1,5-diphosphate carboxylase and glycolate dehydrogenase were determined in these both stages of the division cycle of Chlorella. The results were discussed with reference to the glycollate pathway and the effect of CO 2 concentration during both the growth and the measurement.

Wirkung intermittierender Belichtung auf die 14C-markierten Produkte während der Photosynthese-Induktion von Chlorella vulgaris)

Summary CO2 exchange, 14CO2 fixation and radioactive photosynthetic products of Chlorella vulgaris (strain 211-11 f.) were studied during the induction period at + 25° C using intermittent illumination (10 sec light/20 sec dark). The algae were grown under normal air conditions (0,03 vol.-% CO2) at a temperature of +27° C and a light/dark change of 16:8 h. The transients in CO2 uptake in Chlorella measured with an infrared gas analyzer are characterized at +25° C by exhibition of a maximum and a minimum before reaching steady state of photosynthesis. However under condition of intermittent illumination the «primary CO2 uptake« lasted a relatively long time. The autoradiographic studies of the appearance of labelled products during the induction period showed that under intermittent illumination radioactivity was incorporated into malate and aspartate until 1 minute. 3-phosphoglycerate was labelled only after 1 minute and the intermediates of the Calvin cycle after 3 minutes. On the other hand, at normal illumination conditions 3-phosphoglycerate and the sugar phosphates appeared not only earlier, but were also more strongly labelled. The results show that in Chlorella besides the Calvin cycle a carboxylation of phosphoenolpyruvate exists also at higher temperatures. These two carboxylation pathways are involved in the formation of the transients of photosynthetic CO2 uptake in Chlorella. The results are discussed with reference to the preillumination experiments and the C4 pathway in Chlorella.

Photosynthetic 14CO2 Fixation in Differently Pigmented Anabaena cylindrica

Summary Growth of Anabaena cylindrica at different intensities of white and red light (> 650 nm) was associated with changes in the content and composition of the pigments. The ratio of chlorophyll a to phycocyanin and carotenoids varied from 1:8.1:0.7 in low white light grown cells to 1:6.3:1.3 in cells grown at strong white light and to 1:18.4:0.8 in cultures exposed to red light. The chlorophyll a content was identical in cultures grown in low white and red light and decreased about 50% in those grown in strong white light. The phycocyanin content was enhanced (100%) in Anabaena cells grown in red light and diminished (65%) in those grown in strong white light. Total amino acid content and the values of aspartate, glutamate, glutamine, glycine and arginine studied by high performance liquid chromatography were lower in cells grown in strong white light and red light than in low white light. A markedly enhanced amount of alanine was estimated in strong white light Anabaena extracts. In cells grown in red light, so far not identified primary amines were also detected. Short term kinetics experiments on photosynthetic 14CO2 fixation (percentage proportion of soluble products) of these cyanobacteria cultures showed that the Calvin cycle is the main CO2 assimilation pathway. The percentage proportion of radioactivity of total amino acids arises from low white light to strong white light cells; the main part of the 14C-label could be detected in aspartate, alanine and glycine/serine. A decrease of the 14C-label of sugar monophosphates was found in strong white light cultures in comparison to low white light and red light Anabaena cylindrica. The enzyme activities of ribulose-1,5-bisphosphate carboxylase, aspartate-α-ketoglutarate and aspartate aminotransferase decreased in strong white light grown cells. On the other hand, an increase of activities of malate dehydrogenase (NADPH2 and NADH2), alanine and serine-pyruvate aminotransferases could be observed. Phosphoenolpyruvate carboxylase activity showed practically no change. Our findings indicate that the phosphoenolpyruvate carboxylation reaction must be enhanced in strong white light grown cells.

Light-Dependent Changes in the Lipid and Fatty Acid Composition of Phycocyanin-Free Photosynthetic Lamellae of Synechococcus

Abstract The growth of Synechococcus at different intensities of white and red light caused changes in the pigment composition. The ratio of chlorophyll a to phycocyanin varied from 1:8,2 in LWLI-grown cells to 1:1,4 in cells grown at HWLI and to 1:15,7 in cultures exposed to HRLI. Acyl lipids were quantitatively determ ined and fatty acids of the individual lipid classes analysed by GLC. Phycocyanin-free photosynthetic lam ellae were obtained by fractional centrifugation. No variation was found in the acyl lipid composition of the m em brane preparations. These all contained MGDG, DGDG, SQDG and PG as components. In all the lipids investigated, palmitic, hexadecenoic and octadecenoic acids m ade up to more than 90% of total fatty acids. The pattern of these major components w ithin the lipids from the different cultures depended on the light used. No large differences were detected between zones obtained from LWLI and HRLI isolated membranes, whereas density gradient centrifugation of those from HWLI-grown cells resulted in a completely different pattern of bands. The variations in lipid and fatty acid composition are discussed with respect to changes observed in lipid composition of whole cells and the results reported on tem perature dependent shifts in lipid fluidity in cyanobacteria.