Crucial role of extracellular polysaccharides in desiccation and freezing tolerance in the terrestrial cyanobacterium Nostoc commune

The cyanobacterium Nostoc commune is adapted to the terrestrial environment and has a cosmopolitan distribution. In this study, the role of extracellular polysaccharides (EPS) in the desiccation tolerance of photosynthesis in N. commune was examined. Although photosynthetic O2 evolution was not detected in desiccated colonies, the ability of the cells to evolve O2 rapidly recovered after rehydration. The air-dried colonies contained approximately 10% (wt/wt) water, and field-isolated, natural colonies with EPS were highly water absorbent and were rapidly hydrated by atmospheric moisture. The cells embedded in EPS in Nostoc colonies were highly desiccation tolerant, and O2 evolution was not damaged by air drying. Although N. commune was determined to be a mesophilic cyanobacterium, the cells with EPS were heat tolerant in a desiccated state. EPS could be removed from cells by homogenizing colonies with a blender and filtering with coarse filter paper. This treatment to remove EPS did not damage Nostoc cells or their ability to evolve O2, but O2 evolution was significantly damaged by desiccation treatment of the EPS-depleted cells. Similar to the EPS-depleted cells, the laboratory culture strain KU002 had only small amount of EPS and was highly sensitive to desiccation. In the EPS-depleted cells, O2 evolution was also sensitive to freeze-thaw treatment. These results strongly suggest that EPS of N. commune is crucial for the stress tolerance of photosynthesis during desiccation and during freezing and thawing.     

The role of extracellular polysaccharides produced by the terrestrial cyanobacterium Nostoc sp. strain HK-01 in NaCl tolerance

The terrestrial cyanobacterium Nostoc sp. HK-01 was more tolerant to NaCl stress than the aquatic cyanobacterium Anabaena sp. PCC 7120 (also called Nostoc sp. PCC 7120) which is similar to Nostoc sp. HK-01 in phylogeny. We determined the amount of extracellular polysaccharides (capsular and released polysaccharides) from the cells of both strains cultured with or without 200 mM NaCl. The amount of capsular polysaccharides from Nostoc HK-01 reached approximately 65% of the dry weight whereas that from Anabaena PCC 7120 only occupied approximately 18% of the dry weight under NaCl stress. Anabaena PCC 7120 grew well under NaCl stress when both polysaccharides from Nostoc HK-01 were added to the culture. However, Anabaena PCC 7120 barely grew under NaCl stress when both of its polysaccharides were added. Extracellular polysaccharides from Nostoc HK-01 contained abundant fucose and glucuronic acid in comparison with those from Anabaena PCC 7120. Under NaCl stress, the composition ratios of sugars in the extracellular polysaccharides from Anabaena PCC 7120 hardly changed in comparison with those in ordinary culture conditions. By contrast, the composition ratios of sugars in the extracellular polysaccharides from Nostoc HK-01 changed under NaCl stress. These results suggest that the effect of extracellular polysaccharides from Nostoc HK-01 on NaCl tolerance comes from the increased amount of capsular polysaccharides, the sugar composition, and the change of the sugar composition ratio under NaCl stress.     

Molecular genetic and chemotaxonomic characterization of the terrestrial cyanobacterium Nostoc commune and its neighboring species

The phylogeny of the terrestrial cyanobacterium Nostoc commune and its neighboring Nostoc species was studied using molecular genetic and chemotaxonomic approaches. At least eight genotypes of N. commune were characterized by the differences among 16S rRNA gene sequences and the petH gene encoding ferredoxin-NADP? oxidoreductase and by random amplified polymorphic DNA analysis. The genotypes of N. commune were distributed in Japan without regional specificity. The nrtP gene encoding NrtP-type nitrate/nitrite permease was widely distributed in the genus Nostoc, suggesting that the occurrence of the nrtP gene can be one of the characteristic features that separate cyanobacteria into two groups. The wspA gene encoding a 36-kDa water stress protein was only found in N. commune and Nostoc verrucosum, suggesting that these Nostoc species that form massive colonies with extracellular polysaccharides can be exclusively characterized by the occurrence of the wspA gene. Fifteen species of Nostoc and Anabaena were investigated by comparing their carotenoid composition. Three groups with distinct patterns of carotenoids were related to the phylogenic tree constructed on the basis of 16S rRNA sequences. Nostoc commune and Nostoc punctiforme were clustered in one monophyletic group and characterized by the occurrence of nostoxanthin, canthaxanthin, and myxol glycosides.     

Recovery of photosynthetic systems during rewetting is quite rapid in a terrestrial cyanobacterium, Nostoc commune

Recovery processes of photosynthetic systems during rewetting were studied in detail in a terrestrial, highly drought-tolerant cyanobacterium, Nostoc commune. With absorption of water, the weight of N. commune colony increased in three phases with half-increase times of about 1 min, 2 h and 9 h. Fluorescence intensities of phycobiliproteins and photosystem (PS) I complexes were recovered almost completely within 1 min, suggesting that their functional forms were restored very quickly. Energy transfer from allophycocyanin to the core-membrane linker peptide (L(CM)) was recovered within 1 min, but not that from L(CM) to PSII. PSI activity and cyclic electron flow around PSI recovered within 2 min, while the PSII activity recovered in two phases after a time lag of about 5 min, with half times of about 20 min and 2 h. Photosynthetic CO(2) fixation was restored almost in parallel with the first recovery phase of the PSII reaction center activity. Although the amount of absorbed water became more than 20 times the initial dry weight of the N. commune colony in the presence of sufficient water, about twice the initial dry weight was enough for recovery and maintenance of the PSII activity.     

Protein synthesis and proteolysis in immobilized cells of the cyanobacterium Nostoc commune UTEX 584 exposed to matric water stress.

Cells of the cyanobacterium Nostoc commune UTEX 584 in exponential growth were subjected to acute water stress by immobilizing them on solid supports and drying them at a matric water potential (psi m) of -99.5 MPa. Cells which had been grown in the presence of Na235SO4 before immobilization and rapid drying continued to incorporate 35S into protein for 90 min. This incorporation was inhibited by chloramphenicol. No unique proteins appeared to be synthesized during this time. Upon further drying, the level of incorporation of 35S in protein began to decrease. In contrast, there was an apparent increase in the level of certain phycobiliprotein subunits in solubilized protein extracts of these cells. Extensive proteolysis was detected after prolonged desiccation (17 days) of the cells in the light, although they still remained intact. Phycobilisomes became dissociated in both light- and dark-stored desiccated material.     

Mechanisms to avoid photoinhibition in a desiccation-tolerant cyanobacterium, Nostoc commune

A desiccation-tolerant cyanobacterium, Nostoc commune, showsunique responses to dehydration. These responses are: (i) lossof PSII activity in parallel with the loss of photosynthesis;(ii) loss of PSI activity; and (iii) dissipation of light energyabsorbed by pigment–protein complexes. In this study,the deactivation of PSII is shown to be important in avoidingphotoinhibition when the Calvin–Benson cycle is repressedby dehydration. Furthermore, our evidence suggests that dissipationof light energy absorbed by PSII blocks photoinhibition understrong light in dehydrated states.     

Effect of UV-B radiation on the synthesis of UV-absorbing compounds in a terrestrial cyanobacterium, Nostoc flagelliforme

Effects of two intensities (1 and 5 W?m^?2) of UV-B radiation on the synthesis of UV-absorbing compounds in a terrestrial cyanobacterium Nostoc flagelliforme were investigated. UV-B radiation resulted in lower biomass. Short period (less than 12 h) of UV-B radiation caused an increase of chlorophyll a content, but subsequent duration of treatment (more than 24 h) resulted in a rapid decrease. N. flagelliforme synthesized UV-absorbing compounds such as scytonemin and mycosporine-like amino acids (MAAs) in response to UV-B radiation. Upon 48 h of exposure to UV-B radiation, scytonemin content in cells increased by 103.8 and 164.0 % at 1 and 5 W?m^?2, respectively. Oligosaccharide-linked mycosporine-like amino acids increased by 145.5 % after 12 h at 5 W?m^?2 and 114.5 % after 48 h at 1 W?m^?2 UV-B radiation. HPLC analysis showed that nine MAAs existed in N. flagelliforme cells both from liquid suspension culture and field colony. But the concentration and kinds of them were different. At the two distinct levels of UV-B radiation, the content of particular MAAs increased, declined, or remained unchanged. Moreover, the appearance of two new MAAs was observed.     

Antioxidative activity and chemical constituents of edible terrestrial alga Nostoc commune Vauch

The extract of terrestrial alga Nostoc commune Vauch. has high antioxidative activity. Our study on N. commune Vauch. resulted in the isolation of two β-ionone derivatives, nostocionone and 3-oxo-β-ionone, together with four indole alkaloids, scytonemin, reduced scytonemin, N-(p-coumaroyl)tryptamine, and N-acetyltryptamine. The structures of the isolated compounds were determined on the basis of 1D and 2D NMR and MS analyses. Among these isolates, nostocionone and reduced scytonemin demonstrated strong antioxidative activities which were assessed by using a β-carotene oxidation assay.     

Purification and properties of an enzyme capable of degrading the polysaccharide of the cyanobacterium,Nostoc commune

A novel Nostoc commune-polysaccharide (NPS)-degrading enzyme with a molecular mass of 128.5 kDa was purified from Paenibacillus glycanilyticus DS-1. The optimum pH and temperature of the enzyme activity were 5.5 and 35 degrees C, respectively. The enzyme completely degraded NPS to oligosaccharides, ranging from tetra to hexasaccharides and could degrade the xylan weakly whereas xanthan, gellan, cellulose, curdlan and p-nitrophenyl-beta-D-xylopyranoside were not degraded. Homology analysis of the N-terminal amino acid sequence of the NPS-degrading enzyme against the PIR and SWISS-PROT databases indicated that the sequence was not homologous to any other polysaccharide-degrading enzyme.     

The odd behaviour of carbonic anhydrase in the terrestrial cyanobacterium Nostoc flagelliforme during hydration-dehydration cycles

The terrestrial cyanobacterium Nostoc flagelliforme , inhabiting arid areas, withstands prolonged periods of dehydration. How dehydration and occasional wetting affect inorganic C acquisition in this organism is not well known. As inorganic C acquisition in cyanobacteria often involves carbonic anhydrases (CA), we studied the effect of cycles of hydration and dehydration on the extracellular and intracellular CA activities, at the pH values presumably associated with dew or rain wetting. The external CA of N. flagelliforme (or of the microorganismal consortium of which N. flagelliforme is the main component) is activated by hydration, especially at low pH, and it may facilitate inorganic C acquisition when N. flagelliforme colonies are wetted by dew. Internal CA is present in dry colonies and is rapidly inactivated upon rehydration, therefore an anaplerotic role for this enzyme is proposed.     

Partial characterization of extracellular polysaccharides produced by cyanobacterium Arthrospira platensis

In this study, the physico-chemical characteristics of extracellular polysaccharides (EPS) produced by Arthrospira platensis were evaluated. Elemental analysis and a bicinchoninic acid (BCA) reaction indicated that the EPS were heteropolysaccharides that contain carbohydrate (13%) and protein (55%) moieties. Analysis of the infrared spectrum and elemental analysis revealed the presence of a sulfate group (0.5%). The UV-visible spectrum showed high UV absorption at 190∼230 nm and a shoulder at 260∼280 nm. In addition, this spectrum indicated that EPS can form aggregates with mycosporine-like amino acids and/or scytonemin. Gas chromatography analysis of the carbohydrate portion of the EPS indicated that it was composed of seven neutral sugars: galactose (14.9%), xylose (14.3%), glucose (13.2%), frucose (13.2%), rhamnose (3.7%), arabinose (1%), and mannose (0.3%) and two uronic acids, galacturonic acid (13.5%) and glucuronic acid (0.9%).     

Participation of extracellular polysaccharides in detoxication of cadmium ions by cyanobacteria Nostoc muscorum

Using the methods of absorption and fluorescence spectroscopy, histochemistry and micrography, the influence of cadmium in different concentrations on the exudation of exopolysaccharides by Nostoc muscorum was studied to elucidate the possibility of distance detoxication of cadmium ions by the cyanobacterium with the participation of its exometabolites. The toxic action of Cd was controlled by changes in biomass, trichomes morphology, and the photosynthetic apparatus of the cyanobacterium. The content of exopolysaccharides in the extracellular medium was 0.7 +/- 0.01 gram glucose units per gram dry biomass at the culture density of 2 gram dry biomass per liter. In the initial phase of growth, the exopolysaccharide content decreased to 0.4 +/- 0.1 gram glucose units per gram dry biomass, while upon incubation with cadmium ions it increased several times during a week, after a short adaptation period. The kinetics of exopolysaccharide accumulation and their concentration depended on cadmium content in the medium. After the incubation of N. muscorum with Cd2+, the exometabolites and the mucous coat colored with ditizone, which was indicative of cadmium accumulation. The decomposition of the absorption spectra of the hydrolyzed extracellular metabolites showed that in the presence of cadmium ions, the dominant monosaccharide became glucosamine, which readily binds to Cd2+. It was concluded that the system of N. muscorum protection against cadmium includes the binding of the metal by cell structures, primarily, by the mucilaginous sheath and the distance detoxication by exopolysaccharides in the cultural medium. Cadmium induces the activation of the mucilaginous sheath protective function by changing its composition and the rate of renewal. To provide the distance protection of N. muscorum, the rate of polysaccharide release of the altered primary structure increased.     

Photomophogenesis in the Blue-green Alga Nostoc commune 584

The blue-green alga Nostoc commune 584 displays a photocontrolled developmental cycle similar to that described for N. muscorum A by Lazaroff and Vishniac (1961). In both species white fluorescent light acts at the same stage, ragulating the development of motile trichomes from sheathed aseriate colonies. However white light blocks this step in N. commune 584, whereas the formation of motile trichomes is promoted by white light in N. muscorum A. Light-grown (aseriate) cultures in N. commune 584 were used to determine the action spectra for photomorphogenesis. Green light (max 520 nm) inhbited aseriate colony breakage, and red light (max 640 nm) promoted colony breakage and the differentiation of motile trichomes. On a quantum basis green light was about 3 times more effective than red light. The morphogenetic effects of either red or green light were reversible by irradiation with the other color of light. Repeated photoreversibility was observed, and the algal culutres responded only to the color of the last irradiation in a sequence. An unidentified substance is excreted into the media of motile cultures of both N. commune 584 and N. muscorum A which promotes motility in non-motile cultures. The motility-promoting substances from both species are reciprocally active. Activity is lost when the media are autoclaved.     

Effects of light intensity on components and topographical structures of extracellular polysaccharides from the cyanobacteria Nostoc sp.

A study on the effects of light intensity (40 and 80 μE/m²/sec) on the components and topographical structures of extracellular polysaccharides (EPS) was carried out in cyanobacteria Nostoc sp.. EPS yield increased with light intensity. However, light intensity did not significantly affect the EPS fractions and monosaccharide composition. Higher light intensity generally resulted in higher protein content of EPS in similar fractions. The topographical structure of EPS, investigated by atomic force microscopy, appeared as spherical lumps, chains and networks. The long chains were observed at higher light intensity. Thus, light intensity affected the yield and nature of EPS.     

Cellular differentiation in the cyanobacterium Nostoc punctiforme

Nostoc punctiforme is a phenotypically complex, filamentous, nitrogen-fixing cyanobacterium, whose vegetative cells can mature in four developmental directions. The particular developmental direction is determined by environmental signals. The vegetative cell cycle is maintained when nutrients are sufficient. Limitation for combined nitrogen induces the terminal differentiation of heterocysts, cells specialized for nitrogen fixation in an oxic environment. A number of unique regulatory events and genes have been identified and integrated into a working model of heterocyst differentiation. Phosphate limitation induces the transient differentiation of akinetes, spore-like cells resistant to cold and desiccation. A variety of environmental changes, both positive and negative for growth, induce the transient differentiation of hormogonia, motile filaments that function in dispersal. Initiation of the differentiation of heterocysts, akinetes and hormogonia are hypothesized to depart from the vegetative cell cycle, following separate and distinct events. N. punctiforme also forms nitrogen-fixing symbiotic associations; its plant partners influence the differentiation and behavior of hormogonia and heterocysts. N. punctiforme is genetically tractable and its genome sequence is nearly complete. Thus, the regulatory circuits of three cellular differentiation events and symbiotic interactions of N. punctiforme can be experimentally analyzed by functional genomics.     

Effects of temperature, pH, and UV radiation on alkaline phosphatase activity in the terrestrial cyanobacterium Nostoc flagelliforme

Cyanobacteria produce phosphatases in response to phosphorus deficiency as some other autotrophs. However, little has been documented on the effects of key climate change factors, such as temperature rise and solar UV radiation (280–400 nm), on cyanobacterial alkaline phosphatase activity. Here, we found that the terrestrial cyanobacterium Nostoc flagelliforme showed higher activity of the enzyme with increasing temperature and pH levels, exhibiting maximal values at 45 °C and pH?11, respectively. However, when exposed to solar radiation in the presence of UV-A (320–400 nm) and UV-B (280–320 nm), significant reduction of the enzyme activity was observed at a photosynthetically active radiation (PAR) level of 300 W?m^?2 (1,450 μmol photons m^?2 s^?1), which is equivalent or lower than the noontime level of solar PAR at the organism's habitats. UV-A and UV-A + UV-B induced about 21 and 39 % inhibition of the enzyme activity in the 3-h exposures. The decrease in the activity of phosphatase can be attributed to the UV radiation-induced inactivation of the enzyme and indirectly to the UV radiation-induced production of reactive oxygen species.