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PubMed1996

Driving Forces for Bicarbonate Transport in the Cyanobacterium Synechococcus R-2 (PCC 7942).

Air-grown Synechococcus R-2 (PCC 7942) cultures grown in BG-11 medium are very alkaline (outside pH is 10.0) and use HCO3- as their inorganic carbon source. The cells showed a dependence on Na+ for photosynthesis, but low Na+ conditions (1 mol m-3) were sufficient to support saturating photosynthesis. The intracellular dissolved inorganic carbon in the light was greater than 20 mol m-3 in both low-Na+ conditions and in BG-11 medium containing the usual [Na+] (24 mol m-3, designated high-Na+ conditions). The electrochemical potential for HCO3- in the light was in excess of 25 kJ mol-1, even in high-Na+ conditions. The Na+-motive force was greater than -12 kJ mol-1 under both Na+ conditions. On thermodynamic grounds, an Na+-driven co-port process would need to have a stoichiometry of 2 or greater ([greater than or equal to]2Na+ in/HCO3-1 in), but we show that Na+ or K+ fluxes cannot be linked to HCO3- transport. Na+ and K+ fluxes were unaffected by the presence or absence of dissolved inorganic carbon. In low-Na+ conditions, Na+ fluxes are too low to support the observed net 14C-carbon fixation rate. Active transport of HCO3- hyperpolarizes (not depolarizes) the membrane potential.

Journal Article↗
PubMed1997

Inorganic Carbon Accumulation Stimulates Linear Electron Flow to Artificial Electron Acceptors of Photosystem I in Air-Grown Cells of the Cyanobacterium Synechococcus UTEX 625.

The effect of inorganic carbon (Ci) transport and accumulation on photosynthetic electron transport was studied in air-grown cells of the cyanobacterium Synechococcus UTEX 625. When the cells were depleted of Ci, linear photosynthetic electron flow was almost completely inhibited in the presence of the photosystem I (PSI) acceptor N,N-dimethyl-p-nitrosoaniline (PNDA). The addition of Ci to these cells, in which CO2 fixation was inhibited with glycolaldehyde, greatly stimulated linear electron flow and resulted in increased levels of photochemical quenching and O2 evolution. In aerobic conditions substantial quenching resulted from methyl viologen (MV) addition and further quenching was not observed upon the addition of Ci. In anaerobic conditions MV addition did not result in quenching until Ci was added. Intracellular Ci pools were formed when MV was present in aerobic or anaerobic conditions or PNDA was present in aerobic conditions. There was no inhibitory effect of Ci depletion on electron flow to 2,6-dimethylbenzoquinone and oxidized diaminodurene, which accept electrons from photosystem II. The degree of stimulation of PNDA-dependent O2 evolution varied with the Ci concentration. The extracellular Ci, concentration required for a half-maximum rate (K1/2) was 3.8 [mu]M and the intracellular K1/2 was 1.4 mM for the stimulation of PNDA reduction. These values agreed closely with the K1/2 values of extracellular and intracellular Ci for O2 photoreduction. Linear electron flow to artificial electron acceptors of PSI was enhanced by intracellular Ci, which appeared to exert an effect on PSI or on the intersystem electron transport chain.

Journal Article↗
PubMed1998

Energy sources for HCO3- and CO2 transport in air-grown cells of synechococcus UTEX 625

Light-dependent inorganic C (Ci) transport and accumulation in air-grown cells of Synechococcus UTEX 625 were examined with a mass spectrometer in the presence of inhibitors or artificial electron acceptors of photosynthesis in an attempt to drive CO2 or HCO3- uptake separately by the cyclic or linear electron transport chains. In the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea, the cells were able to accumulate an intracellular Ci pool of 20 mm, even though CO2 fixation was completely inhibited, indicating that cyclic electron flow was involved in the Ci-concentrating mechanism. When 200 m N,N-dimethyl-p-nitrosoaniline was used to drain electrons from ferredoxin, a similar Ci accumulation was observed, suggesting that linear electron flow could support the transport of Ci. When carbonic anhydrase was not present, initial CO2 uptake was greatly reduced and the extracellular [CO2] eventually increased to a level higher than equilibrium, strongly suggesting that CO2 transport was inhibited and that Ci accumulation was the result of active HCO3- transport. With 3-(3,4-dichlorophenyl)-1, 1-dimethylurea-treated cells, Ci transport and accumulation were inhibited by inhibitors of CO2 transport, such as COS and Na2S, whereas Li+, an HCO3--transport inhibitor, had little effect. In the presence of N,N-dimethyl-p-nitrosoaniline, Ci transport and accumulation were not inhibited by COS and Na2S but were inhibited by Li+. These results suggest that CO2 transport is supported by cyclic electron transport and that HCO3- transport is supported by linear electron transport.

Journal Article↗
PubMed1998

Transcriptional and posttranscriptional control of mRNA from lrtA, a light-repressed transcript in Synechococcus sp. PCC 7002.

Transcription regulation and transcript stability of a light-repressed transcript, lrtA, from the cyanobacterium Synechococcus sp. PCC 7002 were studied using ribonuclease protection assays. The transcript for lrtA was not detected in continuously illuminated cells, yet transcript levels increased when cells were placed in the dark. A lag of 20 to 30 min was seen in the accumulation of this transcript after the cells were placed in the dark. Transcript synthesis continued in the dark for 3 h and the transcript levels remained elevated for at least 7 h. The addition of 10 microM rifampicin to illuminated cells before dark adaptation inhibited the transcription of lrtA in the dark. Upon the addition of rifampicin to 3-h dark-adapted cells, lrtA transcript levels remained constant for 30 min and persisted for 3 h. A 3-h half-life was estimated in the dark, whereas a 4-min half-life was observed in the light. Extensive secondary structure was predicted for this transcript within the 5' untranslated region, which is also present in the 5' untranslated region of lrtA from a different cyanobacterium, Synechocystis sp. PCC 6803. Evidence suggests that lrtA transcript stability is not the result of differences in ribonuclease activity from dark to light. Small amounts of lrtA transcript were detected in illuminated cells upon the addition of 25 microg mL-1 chloramphenicol. The addition of chloramphenicol to dark-adapted cells before illumination allowed detection of the lrtA transcript for longer times in the light relative to controls without chloramphenicol. These results suggest that lrtA mRNA processing in the light is different from that in the dark and that protein synthesis is required for light repression of the lrtA transcript.

Bacterial Proteins↗
PubMed1999

Iron superoxide dismutase protects against chilling damage in the cyanobacterium synechococcus species PCC7942

A strain of Synechococcus sp. PCC7942 lacking functional Fe superoxide dismutase (SOD), designated sodB-, was characterized by its growth rate, photosynthetic pigments, inhibition of photosynthetic electron transport activity, and total SOD activity at 0 degrees C, 10 degrees C, 17 degrees C, and 27 degrees C in moderate light. At 27 degrees C, the sodB- and wild-type strains had similar growth rates, chlorophyll and carotenoid contents, and cyclic photosynthetic electron transport activity. The sodB- strain was more sensitive to chilling stress at 17 degrees C than the wild type, indicating a role for FeSOD in protection against photooxidative damage during moderate chilling in light. However, both the wild-type and sodB- strains exhibited similar chilling damage at 0 degrees C and 10 degrees C, indicating that the FeSOD does not provide protection against severe chilling stress in light. Total SOD activity was lower in the sodB- strain than in the wild type at 17 degrees C and 27 degrees C. Total SOD activity decreased with decreasing temperature in both strains but more so in the wild type. Total SOD activity was equal in the two strains when assayed at 0 degrees C.

Journal Article↗
PubMed1999

Protection of photosynthesis against ultraviolet-B radiation by carotenoids in transformants of the cyanobacterium synechococcus PCC7942

The cyanobacterium Synechococcus PCC7942 was transformed with various carotenogenic genes, and the resulting transformants either accumulated higher amounts of beta-carotene and zeaxanthin or showed a shift in the carotenoid pattern toward the formation of zeaxanthin. These transformants were exposed to ultraviolet-B (UV-B) radiation, and the degradation of phycobilins, the inactivation of photosynthetic oxygen evolution, and the activity of photosystem II were determined. In the genetically modified cells, the influence on destruction of phycobilins was negligible. However, protection of photosynthetic reactions against UV-B damage was observed and was dependent on the carotenoid concentrations in the different transformants. Furthermore, it was shown that endogenous zeaxanthin is more effective than beta-carotene. Our results suggest that carotenoids exert their protective function as antioxidants to inactivate UV-B-induced radicals in the photosynthetic membrane.

Journal Article↗
PubMed2000

Inactivation of photosystems I and II in response to osmotic stress in Synechococcus. Contribution of water channels.

The effects of osmotic stress due to sorbitol on the photosynthetic machinery were investigated in the cyanobacterium Synechococcus R-2. Incubation of cells in 1.0 M sorbitol inactivated photosystems I and II and decreased the intracellular solute space by 50%. These effects of sorbitol were reversible: Photosynthetic activity and cytoplasmic volume returned to the original values after removal of the osmotic stress. A blocker of water channels prevented the osmotic-stress-induced inactivation and shrinkage of the intracellular space. It also prevented the recovery of photosynthetic activity and cytoplasmic volume when applied just before release from osmotic stress. Inhibition of protein synthesis by lincomycin had no significant effects on the inactivation and recovery processes, an observation that suggests that protein synthesis was not involved in these processes. Our results suggest that osmotic stress decreased the amount of water in the cytoplasm via the efflux of water through water channels (aquaporins), with resultant increases in intracellular concentrations of ions and a decrease in photosynthetic activity.

Aquaporins↗
PubMed2000

Ionic and osmotic effects of NaCl-induced inactivation of photosystems I and II in Synechococcus sp.

We report here that osmotic effects and ionic effects are both involved in the NaCl-induced inactivation of the photosynthetic machinery in the cyanobacterium Synechococcus sp. PCC 7942. Incubation of the cyanobacterial cells in 0.5 M NaCl induced a rapid and reversible decline and subsequent slow and irreversible loss of the oxygen-evolving activity of photosystem (PS) II and the electron transport activity of PSI. An Na(+)-channel blocker protected both PSII and PSI against the slow, but not the rapid, inactivation. The rapid decline resembled the effect of 1.0 M sorbitol. The presence of both an Na(+)-channel blocker and a water-channel blocker protected PSI and PSII against the short- and long-term effects of NaCl. Salt stress also decreased cytoplasmic volume and this effect was enhanced by the Na(+)-channel blocker. Our observations suggested that NaCl had both osmotic and ionic effects. The osmotic effect decreased the amount of water in the cytosol, rapidly increasing the intracellular concentration of salts. The ionic effect was caused by an influx of Na(+) ions through potassium/Na(+) channels that also increased concentrations of salts in the cytosol and irreversibly inactivated PSI and PSII.

Aquaporins↗
PubMed2001

Unsaturated fatty acids in membrane lipids protect the photosynthetic machinery against salt-induced damage in Synechococcus.

In this study, the tolerance to salt stress of the photosynthetic machinery was examined in relation to the effects of the genetic enhancement of the unsaturation of fatty acids in membrane lipids in wild-type and desA+ cells of Synechococcus sp. PCC 7942. Wild-type cells synthesized saturated and mono-unsaturated fatty acids, whereas desA+ cells, which had been transformed with the desA gene for the Delta12 acyl-lipid desaturase of Synechocystis sp. PCC 6803, also synthesized di-unsaturated fatty acids. Incubation of wild-type and desA+ cells with 0.5 M NaCl resulted in the rapid loss of the activities of photosystem I, photosystem II, and the Na+/H+ antiport system both in light and in darkness. However, desA+ cells were more tolerant to salt stress and osmotic stress than the wild-type cells. The extent of the recovery of the various photosynthetic activities from the effects of 0.5 M NaCl was much greater in desA+ cells than in wild-type cells. The photosystem II activity of thylakoid membranes from desA+ cells was more resistant to 0.5 M NaCl than that of membranes from wild-type cells. These results demonstrated that the genetically engineered increase in unsaturation of fatty acids in membrane lipids significantly enhanced the tolerance of the photosynthetic machinery to salt stress. The enhanced tolerance was due both to the increased resistance of the photosynthetic machinery to the salt-induced damage and to the increased ability of desA+ cells to repair the photosynthetic and Na+/H+ antiport systems.

Cyanobacteria↗
PubMed2001

Nitrogen starvation-induced chlorosis in Synechococcus PCC 7942. Low-level photosynthesis as a mechanism of long-term survival.

Cells of the non-diazotrophic cyanobacterium Synechococcus sp. strain PCC 7942 acclimate to nitrogen deprivation by differentiating into non-pigmented resting cells, which are able to survive prolonged periods of starvation. In this study, the physiological properties of the long-term nitrogen-starved cells are investigated in an attempt to elucidate the mechanisms of maintenance of viability. Preservation of energetic homeostasis is based on a low level of residual photosynthesis; activities of photosystem II and photosystem I were approximately 0.1% of activities of vegetatively growing cells. The low levels of photosystem I activity were measured by a novel colorimetric assay developed from the activity staining of ferredoxin:NADP+ oxidoreductase. Photosystem II reaction centers, as determined by chlorophyll fluorescence measurements, exhibited normal properties, although the efficiency of light harvesting was significantly reduced compared with that of control cells. Long-term chlorotic cells carried out protein synthesis at a very low, but detectable level, as revealed by in vivo [35S]methionine labeling and two-dimensional gel electrophoresis. In conjunction with the very low levels of total cellular protein contents, this implies a continuous protein turnover during chlorosis. Synthesis of components of the photosynthetic apparatus could be detected, whereas factors of the translational machinery were stringently down-regulated. Beyond the massive loss of protein during acclimation to nitrogen deprivation, two proteins that were identified as SomA and SomB accumulated due to an induced expression following nitrogen reduction.

Amino Acid Sequence↗
PubMed1976

Regulatory Properties of the ADP-Glucose Pyrophosphorylase of the Blue-Green Bacterium Synechococcus 6301.

ADP-glucose was found to be the primary sugar nucleotide used for glycogen synthesis by Synechococcus 6301. ADP-glucose pyrophosphorylase was partially purified 12-fold from this blue-green bacterium. The enzyme was activated 8- to 25-fold by glycerate 3-phosphate. Fructose 6-phosphate, fructose 1,6-bisphosphate, 5'-adenylate, and adenosine diphosphate activated the enzyme, but less than glycerate 3-phosphate. The enzyme was inhibited by inorganic phosphate. The I(0.5) of phosphate was 0.072 mm, and in the presence of 2 mm glycerate 3-phosphate, increased to 1.8 mm. The substrate saturation curves for glucose 1-phosphate and ATP were hyperbolic in both the presence and absence of glycerate 3-phosphate or phosphate. The saturation curve for MgCl(2) was sigmoidal; 2 mm glycerate 3-phosphate decreased the sigmoidicity from a Hill slope n value of 5.6 to 2.8, and increased the MgCl(2) optimum from 3 mm to 6 to 7 mm.

Journal Article↗
PubMed1979

Effect of Growth Temperature on the Lipid and Fatty Acid Composition, and the Dependence on Temperature of Light-induced Redox Reactions of Cytochrome f and of Light Energy Redistribution in the Thermophilic Blue-Green Alga Synechococcus lividus.

The thermophilic blue-green alga Synechococcus lividus was grown at 55 and 38 C. Arrhenius plots of the transient reduction of cytochrome during actinic illumination with light that excited both pigment systems revealed breaks near 43 and 26 C for cells grown at 55 C. In cells grown at 38 C these breaks occurred near 37 and 28 C, respectively. The shift from pigment state 1 to state 2 measured by fluorescence transients also showed characteristic breaks in the Arrhenius plots at 44 C for cells grown at 55 C and at 37 to 38 C and possibly at 25 C for cells grown at 38 C. The break points in the Arrhenius plots for the state shift as well as for the cytochrome f reduction are discussed in relation to phase transitions of thylakoid membrane lipids as studied by the temperature dependence of chlorophyll a fluorescence.The variations of fatty acid composition with growth temperature was also studied. When the growth temperature was lowered from 55 to 38 C, the amount of the saturated fatty acid 18:0 in the negatively charged lipids sulfoquinovosyl diglyceride and phosphatidyl glycerol decreased while the unsaturated fatty acids 18:1 and 16:1 increased. In mono- and digalactosyl diglycerides the saturated fatty acids 18:0 and 16:0 decreased and the unsaturated fatty acid 16:1 increased. In general there was an increase in the more fluid lipids in all of the lipid classes when the cells were grown at the lower temperature.

Journal Article↗
PubMed1982

Photosynthesis and Inorganic Carbon Usage by the Marine Cyanobacterium, Synechococcus sp.

The marine cyanobacterium, Synechococcus sp. Nägeli (strain RRIMP N1) changes its affinity for external inorganic carbon used in photosynthesis, depending on the concentration of CO(2) provided during growth. The high affinity for CO(2) + HCO(3) (-) of air-grown cells (K((1/2)) < 80 nanomoles [pH 8.2]) would seem to be the result of the presence of an inducible mechanism which concentrates inorganic carbon (and thus CO(2)) within the cells. Silicone-oil centrifugation experiments indicate that the inorganic carbon concentration inside suitably induced cells may be in excess of 1,000-fold greater than that in the surrounding medium, and that this accumulation is dependent upon light energy. The quantum requirements for O(2) evolution appear to be some 2-fold greater for low CO(2)-grown cells, compared with high CO(2)-grown cells. This presumably is due to the diversion of greater amounts of light energy into inorganic carbon transport in these cells.A number of experimental approaches to the question of whether CO(2) or HCO(3) (-) is primarily utilized by the inorganic carbon transport system in these cells show that in fact both species are capable of acting as substrate. CO(2), however, is more readily taken up when provided at an equivalent concentration to HCO(3) (-). This discovery suggests that the mechanistic basis for the inorganic carbon concentrating system may not be a simple HCO(3) (-) pump as has been suggested. It is clear, however, that during steady-state photosynthesis in seawater equilibrated with air, HCO(3) (-) uptake into the cell is the primary source of internal inorganic carbon.

Journal Article↗
PubMed1984

Salt adaptation of the cyanobacterium synechococcus 6311 growing in a continuous culture (turbidostat).

Physiological aspects of salt-adaptation in the cyanobacterium Synechococcus 6311 growing in a continuous culture (turbidostat) were studied. The process of salt-adaptation was completed within 3 days, as expressed by the specific growth rate of cells grown in the presence of 0.2 and 0.4 molar NaCl. An increase in photosynthetic activity during the adaptation period leads to the accumulation of soluble sugars, essential for osmoregulation in the salt-grown cells. Cells grown in the presence of 0.4 molar NaCl showed an initial inhibition in the rate of protein synthesis which was enhanced after the 1st day of salt stress. After adaptation, salt-grown cells showed an increase in phycobiliprotein synthesis and a higher phycobiliprotein to protein ratio.

Journal Article↗
PubMed1984

Growth and Photosynthesis of the Cyanobacterium Synechococcus leopoliensis in HCO(3)-Limited Chemostats.

Synechococcus leopoliensis was grown in HCO(3) (-)-limited chemostats. Growth at 50% the maximum rate occurred when the inorganic carbon concentration was 10 to 15 micromolar (or 5.6 to 8.4 nanomolar CO(2)). The O(2) to CO(2) ratios during growth were as high as 192,000 to 1. At growth rates below 80% the maximum rate, essentially all the supplied inorganic carbon was converted to organic carbon, and the cells were carbon limited. Carbon-limited cells used HCO(3) (-) rather than CO(2) for growth. They also exhibited a very high photosynthetic affinity for inorganic carbon in short-term experiments. Cells growing at greater than 80% maximum growth rate, in the presence of high dissolved inorganic carbon, were termed carbon sufficient. These cells had photosynthetic affinities that were about 1000-fold lower than HCO(3) (-)-limited cells and also had a reduced capacity for HCO(3) (-) transport. HCO(3) (-)-limited cells are reminiscent of the air-grown cells of batch culture studies while the carbon sufficient cells are reminiscent of high-CO(2) grown cells. However, the low affinity cells of the present study were growing at CO(2) concentrations less than air saturation. This suggests that supranormal levels of CO(2) not required to induce the physiological changes usually ascribed to high CO(2) cells.

Journal Article↗
PubMed1985

A Model for HCO(3) Accumulation and Photosynthesis in the Cyanobacterium Synechococcus sp: Theoretical Predictions and Experimental Observations.

A simple model based on HCO(3) (-) transport has been developed to relate photosynthesis and inorganic carbon fluxes for the marine cyanobacterium, Synechococcus sp. Nägeli (strain RRIMP N1). Predicted relationships between inorganic carbon transport, CO(2) fixation, internal carbonic anhydrase activity, and leakage of CO(2) out of the cell, allow comparisons to be made with experimentally obtained data. Measurements of inorganic carbon fluxes and internal inorganic carbon pool sizes in these cells were made by monitoring time-courses of CO(2) changes (using a mass spectrometer) during light/dark transients. At just saturating CO(2) conditions, total inorganic carbon transport did not exceed net CO(2) fixation by more than 30%. This indicates CO(2) leakage similar to that estimated for C(4) plants.For this leakage rate, the model predicts the cell would need a conductance to CO(2) of around 10(-5) centimeters per second. This is similar to estimates made for the same cells using inorganic carbon pool sizes and CO(2) efflux measurements. The model predicts that carbonic anhydrase is necessary internally to allow a sufficiently fast rate of CO(2) production to prevent a large accumulation of HCO(3) (-). Intact cells show light stimulated carbonic anhydrase activity when assayed using (18)O-labeled CO(2) techniques. This is also supported by low but detectable levels of carbonic anhydrase activity in cell extracts, sufficient to meet the requirements of the model.

Journal Article↗
PubMed1986

Photosynthetic Adaptation by Synechococcus leopoliensis in Response to Exogenous Dissolved Inorganic Carbon.

Synechococcus leopoliensis was grown over a wide range of dissolved inorganic carbon (DIC) concentrations (4-25,000 micromolar) which were obtained by varying culture pH (6.2-9.6) and the CO(2) concentration of the gas stream (36-50,000 microliters per liter). The [DIC] required to half-saturate photosynthesis (K((1/2)) (DIC)) was found to vary depending upon the ambient DIC concentration at which the cells were grown. Low [DIC] grown cells exhibited low values of K((1/2)) (DIC) (4.7 micromolar) whereas cells grown at high [DIC] exhibited high values of K((1/2)) (DIC) (1-2.5 millimolar). Intermediate concentrations of DIC produced intermediate values. Changes in K((1/2)) (DIC) appeared to be solely a function of [DIC] and were independent of both culture pH and CO(2) concentration. As changes in K((1/2)) (DIC) occur in response to DIC concentrations commonly found in natural systems we suggest this adaptation may be of ecological significance.

Journal Article↗
PubMed1987

Na-Stimulation of Photosynthesis in the Cyanobacterium Synechococcus UTEX 625 Grown on High Levels of Inorganic Carbon.

Photosynthesis of washed cells of Synechococcus UTEX 625 grown on 5% CO(2) was markedly stimulated (647 +/- 50%) at pH 8.0 by the addition of low concentrations of NaCl (concentration required for half-maximal response, K((1/2),) = 18 micromolar). Studies with KCl and Na(2)SO(4) showed that the stimulation was due to Na(+). Photosynthesis at pH 6.1 was only slightly stimulated by Na(+). The response of photosynthesis at pH 8.0 to [Na(+)] was strongly sigmoidal for dissolved inorganic carbon ([DIC] </= 500 micromolar). Cells grown with high total [DIC], but air-levels of CO(2), at pH 9.6 showed the same response to low [Na(+)]. The absence of Na(+) could be partially, but not completely overcome, by higher [DIC]. Various methods for examining CO(2) or HCO(3) (-) use (K((1/2)) (CO(2) ) determination; isotopic disequilibrium; and consideration of HCO(3) (-) dehydration rate) were consistent with CO(2) use by the cells, but HCO(3) (-) use could not be ruled out. Isotopic disequilibrium studies showed that CO(2) use was stimulated by Na(+). Cells grown on 5% CO(2) accumulated DIC against a concentration gradient by a process (or processes) dependent on Na(+). No evidence for uptake of Na(+) concomitant with DIC uptake could be found. The lack of O(2) evolution during the initial and most rapid period of DIC accumulation suggested that the required energy was obtained from cyclic photophosphorylation.

Journal Article↗