Modulation of membrane fluidity by catalytic hydrogenation affects the chilling susceptibility of the blue-green alga, Anacystis nidulans

All the changes, i.e. the phase separation temperature of thylakoid lipids, shift in the chilling induced increase of K⁺ permeability and decline in photosynthetic O₂-production, respectively, brought about by temperature acclimation in Anacystis nidulans, can be accomplished by homogeneous catalytic hydrogenation of the fatty acids, as well, using a new water-soluble Pd(II) complex, hitherto unknown in biological applications. Since the thermo-adaptation replaced by proper hydrogenation conducted under isothermal condition results in a similar modification of chilling susceptibility, it afforts direct evidence that chilling response is mediated by changing the degree of fatty acid unsaturation in Anacystis nidulans.     

Synthesis of DNA during the cell cycle of synchronous Anacystis nidulans

Cells of Anacystis nidulans strain 1402-1 incorporate [methyl-³H]thymidine or [8-³H]adenine into DNA; in synchronous cultures (21/2 h full light, 1/2 h weak light, 5 h dark), this incorporation occurs in the dark to different extents according to the labeled precursor offered or to its specific activity. The specific activity of in vivo, uniformly labeled DNA decreases to half the initial value when the cells are grown in the absence of radioactive DNA precursors during the light phase; it does not decrease during the following dark phase. If unlabeled thymidine is given during the dark phase, the specific activity of the DNA starts to decrease at the onset of the next light phase. The time course of the decrease supports the hypothesis that all cells start their DNA replication immediately after illumination and that the first cells have completed if after 1.25 h. The slowest cells then need 3.75 h for completion of DNA replication. It is discussed whether the incorporation during the dark might be due to pool size effects.     

Valyl tRNA's of Anacystis nidulans

Anacystis nidulans was found to contain three tRNA^val isoacceptors which could be charged also in heterologous systems with aminoacyl synth     

Relationship between growth temperature of Anacystis nidulans and phase transition temperature of its thylakoid membranes

The temperatures of the lipid phase transition at which the solid phase disappears were determined by using the X-ray diffraction method in thylakoid membranes of the blue-green alga, Anacystis nidulans. The temperatures were determined as 26 and 16°C for cells grown at 38 and 28°C, respectively.     

Biochemical and biophysical characterization of herbicide-resistant mutants of the unicellular cyanobacterium,Anacystis nidulans R2

Two herbicide-resistant mutants of the unicellular cyanobacterium, Anacystis nidulans R2, were obtained by mutagenesis with N-methyl-N′-nitro-N-nitrosoguanidine. These mutants, A. nidulans R2D1 and R2D2, were selected by growth of mutagenized cells in the presence of 10^?6 M and 10^?5 M 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU), respectively. Both were found to be cross-resistant to 2-chloro-4-ethylamino-6-isopropylamino-s-triazine (atrazine) and 2-n-heptyl-4-hydroxyquinoline-n-oxide (HQNO) by measurement of Photosystem II activity in the presence of the inhibitors. The DCMU-resistance trait from each mutant was transferred to a wild-type genetic background by DNA-mediated transformation of A. nidulans cells. The two resulting transformants, A. nidulans R2D1-X1 and R2D2-X1, were similar to the original mutants with respect to DCMU- and HQNO-resistance. However, both exhibited increased sensitivity to atrazine relative to the mutants from which they were derived. Polyacrylamide gel electrophoretic analysis revealed that the mutants and transformants were deficient in a 34 kDa, surface-exposed polypeptide which was present in the wild-type strain; the transformants exhibited a new polypeptide of 35.5 kDa which was also highly surface-exposed.     

Reversible inhibition of photochemistry of photosystem II by Ca2+ removal from intact cells of Anacystis nidulans

Depletion of Ca²⁺ from Anacystis nidulans produces an inhibition of O₂ evolution that is accompanied both at 39°C and 77 K by a loss of chlorophyll fluorescence of variable yield. This indicates that Ca²⁺-depletion causes disruption of normal photosystem II function, manifested by the disappearance of photoreduction of Q. Delayed light emission in the ms time range is also eliminated in Ca²⁺-depleted cells, which confirms that Ca²⁺ removal prevents charge separation and recombination in reaction centers of photosystem II. Readdition of Ca²⁺ to depleted cells restores fully the fluorescence of variable yield and delayed light emission, as well as O₂ evolution. Thus, Ca²⁺ may be a required component for photosystem II in A. nidulans.     

Primary Role of the Cytoplasmic Membrane in Thermal Acclimation Evidenced in Nitrate-Starved Cells of the Blue-Green Alga, Anacystis nidulans

The lipid phase transition of the cytoplasmic membrane and the chilling susceptibility were studied in nitrate-starved Anacystis nidulans cells. Nitrate starvation resulted in the disappearance of the thylakoid membrane system, without any effect on chilling susceptibility. The chilling susceptibility of the algal cells depended on the growth temperature. Temperatures of lipid phase transitions of the cytoplasmic membranes were detected by chilling-induced spectral changes in the carotenoid region, in vivo. These values were identical to those of cultures containing intact thylakoid systems. Our results suggest that cytoplasmic membrane plays a determinative role in the thermal acclimation of the alga cells.     

Photosynthetic activity of diimidoester-modified cells,permeaplasts, and cell-free membrane fragments of the blue-green alga Anacystis nidulans

On treating the blue-green alga Anacystis nidulans with dimethylsuberimidate up to 70% of the free NH₂ of the photosynthetic membrane is amidinated, and presumably inter- and intramolecular cross-links are established in the membrane proteins. Amidination destroys the ability of A. nidulans to photoreduce HCO₃^? but leaves the photochemical activities of Photosystems II and I nearly intact. With added electron acceptors, photosynthetic O₂ evolution can be demonstrated both with permeable cells (permeaplasts) prepared by digestion of the cell wall of dimethylsuberimidate-reacted A. nidulans with lysozyme, as well as with heavy membrane particles (36 000 × g) prepared from dimethylsuberimidate-reacted cells.Permeaplasts prepared from dimethylsuberimidate-reacted cells resist damage in hypoosmotic medium, whereas those prepared from unreacted cells are induced to release C-phycocyanin. On the other hand, the former are inactivated more easily by heat stress than the latter. On this basis, it is concluded that cross-linking with dimethylsuberimidate confers functional instability to photosynthetic membranes.     

Particle aggregation in photosynthetic membranes of the blue-green alga Anacystis nidulans

Freeze-fracture electron microscopy demonstrates that in photosynthetic membranes of the blue-green alga Anacystis nidulans quenched from a temperature below growth temperature, areas devoid of membrane particles occur. We suggest that this phenomenon is related to phase transitions in the photosynthetic membrane.     

pH-jump-induced phosphorylation of ADP in the cyanobacterium Anacystis nidulans

The low ATP levels in dark anaerobic cells of the cyanobacterium Anacystis nidulans more than doubled within 5 s after rapid addition of HCl shifting external pH from 9.0 to 4.5. Steady-state levels of ATP and intracellular pH remained constant at 0.95 ± 0.15 nmol/mg dry weight and 6.9 ± 0.3, respectively. ΔpH-induced ATP synthesis was inhibited by dicyclohexylcarbodiimide and carbonyl cyanide m-chlorophenylhydrazone but not by carbon monoxide. According to our results the cytoplasmic membrane of A. nidulans has to be regarded as an energy-transducing membrane bioenergetically similar to the thylakoid membrane.     

Ultraviolet Light Inactivation and Photoreactivation of AS-1 Cyanophage in Anacystis nidulans

Black light effected photorecovery of AS-1 cyanophage and wild-type cells. However, only partial photoreactivation of AS-1 was observed in a partially photoreactivable mutant of Anacystis nidulans.     

Preparation of [U-14C]-labelled glycogen, maltosaccharides, maltose, and D-glucose by photoassimilation of 14CO2 in Anacystis nidulans and selective enzymic degradation.

A procedure is described for the isolation from the phototrophic procaryole Anacystis nidulans of [U-¹⁴C]-labelled glycogen, with high specific radioactivity,formed when NaH¹⁴CO₃ was added to non-dividing cells that continued to photoassimilate CO₂. [U-¹⁴C]-Labelled glycogen was then treated with isoamylase (EC 3.2.1.68), isoamylase plus beta-amylase (EC 3.2.1.2), or glucoamylase (EC 3.2.1.3) to give [U-¹⁴C]-labelled maltosaccharides, maltose-U-¹⁴C, or d-glucose-U-¹⁴C, respectively.     

Characterization of the Proton-Translocating Cytochrome c Oxidase Activity in the Plasma Membrane of Intact Anacystis nidulans Spheroplasts

Intact spheroplasts of the cyanobacterium (blue-green alga) Anacystis nidulans oxidized various exogenous c-type cytochromes with concomitant outward proton translocation while exogenous ferricytochrome c was not reduced. The H⁺/e⁻ stoichiometry was close to 1 with each of the cytochromes and did not depend on the actual rate of the oxidase reaction. Observed proton ejections were abolished by the uncoupler carbonyl cyanide m-chlorophenylhydrazone. Cyanide, azide, and carbon monoxide inhibited cytochrome c oxidation and proton extrusion in parallel while dicyclohexylcarbodiimide affected proton translocation more strongly than cytochrome c oxidation. The cytoplasmic membrane of A. nidulans appears to contain a proton-translocating cytochrome c oxidase similar to the one described for mitochondria.     

pH Changes in the Cytoplasm of the Blue-Green Alga Anacystis nidulans Caused by Light-dependent Proton Flux into the Thylakoid Space

The pH in the cytoplasmic and thylakoid spaces of the blue-green alga, Anacystis nidulans, has been determined in the light and in the dark by uptake of 5,5-dimethyloxazolidine-2,4-dione and methylamine into the sucrose-impermeable ³H-H₂O space, as measured by silicon layer filtering centrifugation.     

Characterization of a ribonuclease from Anacystis nidulans infected with cyanophage as-1

In Anacystis nidulans the ribonuclease (RNase) activity is very low but is greatly increased upon phage-infection. A RNase was isolated and purified over 300-fold from A. nidulans cells infected by cyanophage AS-1. The enzyme did not attack single- or double-stranded DNA, was inactive on p-nitrophenyl phosphate or bis-p-nitrophenyl phosphate as substrates, and had neither 3′- nor 5′-nucleotidase activity. The approximate MW of the enzyme was 12000. Maximal enzyme activity was at pH 7.5. No absolute requirement for metal ions was observed, but Fe³⁺ stimulated and Co²⁺ and Ni²⁺ inhibited enzyme activity. The enzyme is an endonuclease which, upon exhaustive hydrolysis, produces mainly oligonucleotides (average chain-length: 3) with 3′-P termini. Analysis of the base composition of these oligonucleotides and determination of their 3′-terminal nucleosides, together with the investigation of the rate of hydrolysis of synthetic polyribonucleotides, have shown that the enzyme has a relative specificity for uridylic acid.     

Characterization of Sulphate Uptake in Anacystis nidulans

Sulphate uptake in the blue-green alga Anacystis nidulans appears based upon an active mechanism with a Km of 0.75 μM and V_max of 0.7 pmol/min × 10⁶ cells. Sulphate uptake is competitively inhibited by thiosulphate and sulphite. The sulphate uptake has a pH optimum at 8 and a temperature optimum at 40°C. By increasing the extracellular sulphate concentration from 0.1 to 10 μM the sulphate pool in Anacystis was altered from 8.3, 10^?5M to 5.9, 10^?4M.     

Induction of temperate cyanophage AS-1 by heavy metal - copper

Background  

It has been reported that some marine cyanophage are temperate and can be induced from a lysogenic phase to a lytic phase by different agents such as heavy metals. However, to date no significant reports have focused on the temperate nature of freshwater cyanophage/cyanobacteria. Previous experiments with cyanophage AS-1 and cyanobacteria Anacystis nidulans have provided some evidence that AS-1 may have a lysogenic life cycle in addition to the characterized lytic cycle.     

Influence of Iron Deprivation on the Membrane Composition of Anacystis nidulans

Cultures of the cyanobacterium Anacystis nidulans were grown under iron-deficient conditions and then restored by the addition of iron. Membrane proteins from iron-deficient and iron-restored cells were analyzed by lithium dodecyl sulfate-polyacrylamide gradient gel electrophoresis. The incorporation of [³⁵S]sulfate into membrane proteins and lactoperoxidase-catalyzed ¹²⁵I iodination were used to monitor the rates of polypeptide biosynthesis and surface exposure of membrane proteins, respectively. These polypeptide profiles revealed major differences in the membrane composition of iron-deficient and normal cells. Iron deficiency caused a decrease in the amount of certain important membrane proteins, reflecting a decreased rate of biosynthesis of these peptides. Several photosystem II peptides also showed an increase in surface exposure after iron stress. In addition, iron deficiency led to the synthesis of proteins at 34 and 52 kilodaltons which were not present in normal cells. When iron was restored to a deficient culture, a metabolic sequence was initiated within the first 12 h after the addition of iron which led to phenotypically normal cells. Pulse labeling with [³⁵S]sulfate during this period demonstrated that iron addition initiates a coordinated pattern of synthesis that leads to the assembly of normal membranes.     

Relationships between the Transition of the Physical Phase of Membrane Lipids and Photosynthetic Parameters in Anacystis nidulans and Lettuce and Spinach Chloroplasts

The transition of the physical phase of lipids in membrane fragments of a blue-green alga Anacystis nidulans was studied by a spin labeling technique. The maximum hyperfine splitting of the electron spin resonance spectrum of the N-oxyl-4′, 4′-dimethyloxazolidine derivative of 5-ketostearic acid plotted against the reciprocal of the absolute temperature gave a discontinuity point that was characteristic of a transition of the physical phase of the hydrocarbon region of membrane lipids. The phase transition appeared at approximately 13 or 24 C in the organisms grown at 28 or 38 C, respectively.     

Active sodium extrusion reduces net efficiencies of oxidative phosphorylation in the strictly photoautotrophic cyanobacterium Anacystis nidulans

Efficiencies of oxidative phosphorylation ($\text{P}\text{O}$ ratios), intracellular high-energy phosphate pools (ATP and ADP) under aerobic and anaerobic dark conditions, and photosynthetic oxygen evolution measured with intact cells of Anacystis nidulans were found to be specifically depressed by NaCl, but not by KCl. A scheme is proposed which explains the deleterious effect of sodium on the energy metabolism of A. nidulans by competition for protons between ATP synthesis and active sodium extrusion.