Photomovement of the Gliding Cyanobacterium Synechocystis sp. PCC 6803

Abstract— Using a computerized videomicroscope motion analysis system, we investigated the photomovements of two Synechocystis sp. (PCC 6803 and ATCC 27184). Synechocystis sp. PCC 6803 displays a relatively slow gliding motion. The phototactic and photokinetic speeds of this cyanobacterium in liquid media were 5μm/min and 15.8 μm/min, respectively, at 3μmol/m²/s of stimulant white light. Synechocystis sp. PCC 6803 senses light direction rather than intensity for phototaxis. Synechocystis sp. ATCC 27184 showed a weak photokinesis but no phototaxis. Analysis of Synechocystis sp. ATCC 27184 suggests that the loss of phototaxis results from spontaneous mutation during several years of subculture. When directional irradiation was applied, the cell population of Synechocystis sp. PCC 6803 began to deviate from random movement and reached maximum orientation at 5 min after the onset of stimulant white light. Synechocystis sp. PCC 6803 showed high sensitivity to the stimulant white light of fluence rates as low as 0.002 |unol/m²/s. Neither 1,3-dichlorophenyldimethyl urea nor cyanide affected phototactic orientation, whereas cyanide inhibited gUding speed. This result suggests that the phototaxis of Synechocystis sp. PCC 6803 is independent of photosynthetic phosphorylation and that its gliding movement is primarily powered by oxidative phosphorylation. In the visible wavelength region, 560 nm, 660 nm and even 760 nm caused positive phototaxis. However, 360 nm light induced strikingly negative phototaxis. Therefore, at least two independent photoreceptors may exist to control phototaxis. The photoreceptor for positive phototaxis appears likely to be a phytochrome-like tetrapyrrole rather than chlorophyll a .     

Induction of mycosporine-like amino acids (MAAs) in cyanobacteria by solar ultraviolet-B radiation

Three filamentous and heterocystous N₂-fixing cyanobacteria, Anabaena sp., Nostoc commune and Scytonema sp. were tested for the presence of ultraviolet-absorbing mycosporine-like amino acids (MAAs) and their induction by solar ultraviolet-B (UV-B) radiation. High performance liquid chromatographic (HPLC) studies revealed the presence of only one type of MAAs in all three cyanobacteria, that was identified as shinorine, a bisubstituted MAA containing both glycine and serine groups having an absorption maximum at 334 nm and a retention time of around 2.8 min. There was a circadian induction in the synthesis of MAAs when the cultures were exposed to mid-latitude solar radiation (Playa Unión, Rawson, Chubut, Patagonia, Argentina) for 3 days, 4–6th February, 2000. Solar radiation was measured by an ELDONET (European Light Dosimeter Network) filter radiometer permanently installed on the roof of the Estación de Fotobiología Playa Unión (43°18′ S; 65°03′ W). The maximum irradiances were around 450–500, 45–50 and 1.0–1.2 W m⁻² for PAR (photosynthetic active radiation), UV-A (ultraviolet-A) and UV-B (ultraviolet-B), respectively. PAR and UV-A had no significant impact on MAA induction while UV-B induced the synthesis of shinorine in all three cyanobacteria. Shinorine was found to be induced mostly during the light period. During the dark period the concentration stayed almost constant. In addition to shinorine, another unidentified, water-soluble, brownish compound with an absorption maximum at 315 nm was found to be induced by UV-B only in Scytonema sp. and released into the medium. This substance was neither found in Anabaena sp. nor in Nostoc commune. Judging from the results, the studied cyanobacteria may protect themselves from deleterious short wavelength radiation by their ability to synthesize photoprotective compounds in response to UV-B radiation.     

Photoprotection and long-term acclimation to UV radiation in the marine diatom Thalassiosira weissflogii.

Unialgal cultures of the marine diatom Thalassiosira weissflogii (Grunow) were exposed for 40 days to artificial UV-B radiation in the presence of PAR and UV-A to examine long-term acclimation to UV, PAR and UV-A were supplied 14 h daily, while UV-B (two levels: 0.16 and 0.30 W m(-2) unweighted) was supplied for 4 h/day. Growth rates and photochemical capacity (CFC ratio) both decreased over the first 10-15 days, then recovered. No obvious differences were noted between the responses to the two UV-B treatments. The concentration of the major pigments (chlorophyll a and c(1+2), fucoxanthin and beta,beta-carotene) changed very little with time, except for diatoxanthin. which increased over the first 16 days, decreased over the next 13 days, then increased again from day 29 to the end of the experiment. The concentration of total mycosporine-like amino acids (MAAs) was initially undetectable, then increased from day 16 in the high UV-B treatment and after day 22 in the low UV-B treatment, reaching a maximum on day 29 for both treatments and decreasing afterwards. The synthesis of MAAs proceeded only once photochemical capacity had recovered from the initial UV stress and this recovery likely involved the xanthophyll cycle (diatoxanthin increase). The concentration of MAAs decreased when the cells showed signs of photoinhibition (decrease in CFC ratio). It also showed an inverse trend with diatoxanthin. UV-B alone had little regulatory effect over these responses, except possibly for an earlier synthesis of MAAs under HUV-B conditions. This suggests that the observed changes were due to UV-A rather than to UV-B exposure. The overall response of this coastal diatom to prolonged UV exposure indicates that T. weissflogii is a relatively UV-tolerant species and that its long-term response to UV exposure involves an activation of the xanthophyll cycle followed by the synthesis of MAAs, which may proceed only when photoinhibition is relieved.     

Exogenous Spermidine Alleviates UV-Induced Growth Inhibition of Synechocystis sp. PCC 6803 via Reduction of Hydrogen Peroxide and Malonaldehyde Levels

Effects of exogenously added spermidine (Spd) to UV-treated Synechocystis sp. PCC 6803 cultures on their growth, intracellular pigments, hydrogen peroxide (H₂O₂), malonaldehyde (MDA) contents, and antioxidant enzymes were investigated. Growth inhibition of cells subjected to 1-h UV-A, UV-B, and UV-C irradiation was abolished in culture added with 0.5 mM Spd. Both chlorophyll a and carotenoid contents were decreased under UV radiations in cells grown in BG₁₁ medium. However, the contents of these two pigments were slightly increased under UV radiations in Spd-supplemented cells with the consequence of enhanced oxygen evolution. Intracellular levels of H₂O₂ and MDA generated during 1-h UV irradiation were decreased when the culture medium contained 0.5 mM Spd. The antioxidative enzymes, catalase, and superoxide dismutase had a little or no response towards Spd supplementation under UV irradiation except for some increase in superoxide dismutase activity under UV-C. Total intracellular polyamines were decreased during Spd supplementation under UV stress; however, the cells showed a drastic increase in the amount of Put under this condition. Altogether, exogenous Spd is likely a potential compound that enables Synechocystis cells to cope with UV stress.     

Effect of UV-C on Algal Evolution and Differences in Growth Rate, Pigmentation and Photosynthesis Between Prokaryotic and Eukaryotic Algae

Insight into the influence of UV-C radiation on the evolutionary relationship between prokaryotic and eukaryotic algae was studied in seven species of algae exposed to different UV-C irradiances. The order of their acclimation (from most tolerant to sensitive) is Synechococcus sp. PCC7942 (Cyanophyta), Synechocystis sp. PCC6803 (Cyanophyta), Chlorella protothecoides (Chlorophyta), Chlamydomonas reinhardtii (Chlorophyta), Phaeodactylum  tricornutum (Bacillariophyta), Alexandrium  tamarense (Pyrrhophyta) and Dicrateria  zhanjiangensis (Chrysophyta). These results are in accordance with the algal evolution process that is generally accepted and proved by fossil record. It shows that UV-C radiation is an important environmental factor that cannot be ignored in the evolutionary process from prokaryotic algae to eukaryotic algae. The threshold of UV-C radiation at which prokaryotic algae can survive but eukaryotic algae cannot was found to be approximately 0.09 W m⁻². This was the first time to determine with precision the irradiance level at which UV-C contributed as a selection pressure of evolution. Furthermore, the effects of UV-C radiation on photosynthetic performance, growth rate and pigment content were investigated in two species of prokaryotic algae: Synechococcus sp. PCC7942 and Synechocystis sp. PCC6803, and two species of eukaryotic algae: C. reinhardtii and C. protothecoides . After 6 days of exposure, the contents of chlorophyll a and carotenoids decreased in all species, moreover reduction in C. reinhardtii and C. protothecoides was more obvious than in Synechococcus sp. PCC7942 and Synechocystis sp. PCC6803. The ability to photosynthesize followed the same trend as the pigments.     

Variations in Photosystem I Properties in the Primordial Cyanobacterium Gloeobacter violaceus PCC 7421

We compared the optical properties of the trimeric photosystem (PS) I complexes of the primordial cyanobacterium Gloeobacter violaceus PCC 7421 with those of Synechocystis sp. PCC 6803. Gloeobacter violaceus PS I showed (1) a shorter difference maximum of P700 by approximately 2 nm, (2) a smaller antenna size by approximately 10 chlorophyll (Chl) a molecules and (3) an absence of Red Chls. The energy transfer kinetics in the antennae at physiological temperatures were very similar between the two species due to the thermal equilibrium within the antenna; however, they differed at 77 K where energy transfer to Red Chls was clearly observed in Synechocystis sp. PCC 6803. Taken together with the lower P700 redox potential in G. violaceus by approximately 60 mV, we discuss differences in the optical properties of the PS I complexes with respect to the amino acid sequences of core proteins and further to evolution of cyanobacteria.     

Red antenna states of photosystem I from cyanobacteria Synechocystis PCC 6803 and Thermosynechococcus elongatus: single-complex spectroscopy and spectral hole-burning study

Hole-burning and single photosynthetic complex spectroscopy were used to study the excitonic structure and excitation energy-transfer processes of cyanobacterial trimeric Photosystem I (PS I) complexes from Synechocystis PCC 6803 and Thermosynechococcus elongatus at low temperatures. It was shown that individual PS I complexes of Synechocystis PCC 6803 (which have two red antenna states, i.e., C706 and C714) reveal only a broad structureless fluorescence band with a maximum near 720 nm, indicating strong electron-phonon coupling for the lowest energy C714 red state. The absence of zero-phonon lines (ZPLs) belonging to the C706 red state in the emission spectra of individual PS I complexes from Synechocystis PCC 6803 suggests that the C706 and C714 red antenna states of Synechocystis PCC 6803 are connected by efficient energy transfer with a characteristic transfer time of approximately 5 ps. This finding is in agreement with spectral hole-burning data obtained for bulk samples of Synechocystis PCC 6803. The importance of comparing the results of ensemble (spectral hole burning) and single-complex measurements was demonstrated. The presence of narrow ZPLs near 710 nm in addition to the broad fluorescence band at approximately 730 nm in Thermosynechococcus elongatus (Jelezko et al. J. Phys. Chem. B 2000, 104, 8093-8096) has been confirmed. We also demonstrate that high-quality samples obtained by dissolving crystals of PS I of Thermosynechococcus elongatus exhibit stronger absorption in the red antenna region than any samples studied so far by us and other groups.     

Effect of UV-B Radiation and Desiccation Stress on Photoprotective Compounds Accumulation in Marine <Emphasis Type="Italic">Leptolyngbya</Emphasis> sp.

Increased awareness regarding the harmful effects of ultraviolet (UV)-B radiation has led to the search for new sources of natural UV-B protecting compounds. Mycosporine-like amino acids are one of such promising compounds found in several organisms. Cyanobacteria are ideal organisms for isolation of these compounds due to their compatibility and adaptability to thrive under harsh environmental conditions. In the following investigation, we report the production of shinorine in Leptolyngbya sp. isolated from the intertidal region. Based on the spectral characteristics and liquid chromatography-mass spectrometry analysis, the UV-absorbing compound was identified as shinorine. To the best of our knowledge, this is the first report on the occurrence of shinorine in Leptolyngbya sp. We also investigated the effect of artificial UV-B radiation and periodic desiccation on chlorophyll-a, total carotenoids, and mycosporine-like amino acids (MAAs) production. The UV-B radiation had a negative effect on growth and chlorophyll concentration, whereas it showed an inductive effect on the production of total carotenoids and MAAs. Desiccation along with UV-B radiation led to an increase in the concentration of photoprotective compounds. These results indicate that carotenoids and MAAs thus facilitate cyanobacteria to avoid and protect themselves from the deleterious effects of UV-B and desiccation.     

Light-generated paramagnetic intermediates in BLUF domains

Blue-light sensitive photoreceptory BLUF domains are flavoproteins, which regulate various, mostly stress-related processes in bacteria and eukaryotes. The photoreactivity of the flavin adenine dinucleotide (FAD) cofactor in three BLUF domains from Rhodobacter sphaeroides, Synechocystis sp. PCC 6803 and Escherichia coli have been studied at low temperature using time-resolved electron paramagnetic resonance. Photoinduced flavin triplet states and radical-pair species have been detected on a microsecond time scale. Differences in the electronic structures of the FAD cofactors as reflected by altered zero-field splitting parameters of the triplet states could be correlated with changes in the amino-acid composition of the various BLUF domains' cofactor binding pockets. For the generation of the light-induced, spin-correlated radical-pair species in the BLUF domain from Synechocystis sp. PCC 6803, a tyrosine residue near the flavin's isoalloxazine moiety plays a critical role.     

UV-B-induced synthesis of mycosporine-like amino acids in three strains of Nodularia (cyanobacteria)

Three filamentous and heterocystous cyanobacterial strains of Nodularia, Nodularia baltica, Nodularia harveyana and Nodularia spumigena, have been tested for the presence and induction of ultraviolet-absorbing/screening mycosporine-like amino acids (MAAs) by simulated solar radiation in combination with 395 (receiving photosynthetically active radiation (PAR) only), 320 (receiving PAR + UV-A) and 295 (receiving PAR + UV-A + UV-B) nm cut-off filters. Absorption spectroscopic analyses of the methanolic extracts of samples revealed a typical MAA peak at 334 nm in all three cyanobacteria. Specific contents of MAAs had a pronounced induction in the samples covered with 295 nm cut-off filters after 72 h of irradiation. In comparison, there was little induction of MAAs in the samples covered by 395 and 320 nm cut-off filters. High performance liquid chromatographic (HPLC) studies revealed the presence of two types of MAAs in all three cyanobacteria, which were identified as shinorine and porphyra-334, both absorbing maximally at 334 nm. The occurrence of porphyra-334 is rare in cyanobacteria. Specific content of both shinorine and porphyra-334 were induced remarkably only in the samples covered with 295 nm cut-off filters. The results indicate that in comparison to UV-A and PAR, UV-B is more effective in eliciting MAAs induction in the studied cyanobacteria.     

Long-lived coherent oscillations of the femtosecond transients in cyanobacterial photosystem I

The pulsed excitation of electronic levels coupled to specific nuclear modes by a 26 fs laser pulse at 706 nm creates a wavepacket in the nuclear space of photopystem I (PS I) of Synechocystis sp. strain PCC 6803 both in the ground state and in the one-exciton manifold. Fourier transform of transient decay curves shows several low frequency peaks. The most prominent Power Spectral Density (PSD) peaks are at omega = 49 cm(-1) and omega = 88 cm(-1). The peculiarity of the coherent wavepacket in the PS I of S. sp. strain PCC 6803 is the unique, long-lived 49 cm(-1) and 88 cm(-1) oscillations with decay times up to 10 ps. It was suggested that such a long-lived coherence is determined by a contribution of the ground state wavepacket. The dependence of these two PSD peaks on the probe wavelength resembles the profile of the transient absorption spectra of PS I. The pump-probe signal in the Soret region reflects the dynamics of the ground state wavepacket created by pulsed excitation of the Q(y)-band. It was shown that the multimode Brownian oscillator model allows a quantitative fit of the oscillatory patterns of the pump-probe signal to be obtained.     

Influence of PAR and UV-A in determining plant sensitivity and photomorphogenic responses to UV-B radiation

The role of photosynthetically active radiation (400-700 nm) (PAR) in modifying plant sensitivity and photomorphogenic responses to ultraviolet-B (280-320 nm) (UV-B) radiation has been examined by a number of investigators, but few studies have been conducted on ultraviolet-A (320-400 nm) (UV-A), UV-B and PAR interactions. High ratios of PAR-UV-B and UV-A-UV-B have been found to be important in ameliorating UV-B damage in both terrestrial and aquatic plants. Growth chamber and greenhouse studies conducted at low PAR, low UV-A and high UV-B often show exaggerated UV-B damage. Spectral balance of PAR, UV-A and UV-B has also been shown to be important in determining plant sensitivity in field studies. In general, one observes a reduction in total biomass and plant height with decreasing PAR and increasing UV-B. The protective effects of high PAR against elevated UV-B may also be indirect, by increasing leaf thickness and the concentration of flavonoids and other phenolic compounds known to be important in UV screening. The quality of PAR is also important, with blue light, together with UV-A radiation, playing a key role in photorepair of DNA lesions. Further studies are needed to determine the interactions of UV-A, UV-B and PAR.     

EFFECTS OF UV IRRADIATION ON A LIVING SKIN EQUIVALENT

Abstract— The Living Skin Equivalent (LSE™) is an organotypic coculture composed of human dermal fibroblasts interspersed in a collagen-containing matrix and overlaid with human keratinocytes forming a stratified epidermis. The LSE has a dry, air-exposed epidermal surface suitable for the application of oils, creams and emulsions. These features suggested its feasibility as an in vitro skin model for studying the protective effects of sunscreens. Using the thiazolyl blue (MTT) conversion assay as a measure of mitochondrial function, the extent of cytotoxicity induced by various doses of UV-R (280–400 nm) or UV-A (320–400 nm) was evaluated in the LSE. The doses of UV radiation that caused 50% reductions in MTT conversion (UV-R₅₀ or UV-A₅₀) in different lots of LSE were 0.053 ± 0.021 J/cm² (n = 29) and 11.6 ± 4.9 J/cm² (n = 17) for UV-R and UV-A, respectively. The protective effects of an 8% homosylate standard and of five UV-A sunscreens, topically applied to the LSE, were determined and compared with their reported protection factors in human skin. Morphological changes and the release of proinflammatory mediators (interleukin-1-α, tumor necrosis factor-α and prostaglandin E2) implicated in UV-induced erythema were also demonstrated in the LSE exposed to UV-A or UV-B. The data suggest that the LSE can be used for studying the effects of U V radiation on skin and may have utility for assessing the efficacy of certain sunscreens against UV-B and UV-A.     

Characterization of 1,4-dihydroxy-2-naphthoyl-coenzyme A synthase (MenB) in phylloquinone biosynthesis of Synechocystis sp. PCC 6803

The gene product Sll1127 is a predicted 1,4-dihydroxy-2-naphthoyl-CoA synthase catalyzing an intramolecular Claisen condensation in the phylloquinone biosynthesis of the cyanobacterium Synechocystis sp.PCC 6803.This predicted catalytic function has been verified and the enzyme has been characterized for the first time with kcat = 0.013 s-1 and KM = 9μM.Its catalytic activity is found to strictly depend on externally added bicarbonate with an apparent KD = 0.60 mM.In addition,this enzyme is inhibited by its 1,4-dihydroxy-2-naphthoyl-CoA product through high-affinity binding,which causes a 18 nm shift of the inhibitor absorption at 392 to 410 nm and engenders a new absorption peak at 345 nm.All these properties of the cyanobacterial enzyme are closely similar to those of the Escherichia coli orthologue from the menaquinone biosynthetic pathway.These results provide additional supporting evidence for the essential role of bicarbonate as a catalytic base in the enzymatic reaction and the eubacterial origin of the enzymes in the cyanobacterial biosynthesis of phylloquinone.     

UV-ABC screens of luteolin derivatives compared to edelweiss extract

Pure luteolin is a remarkably heat (200°C/6 days) and UV stable UV-A screen, however, native luteolin enriched to 37% in an edelweiss extract lost its UV-A screen properties upon UV irradiation (～4MJm(-2)). This contrasting behavior led to the examination of a series of purified luteolin derivatives as UV screen candidates. 3',4',5,7-Tetralipoyloxyflavones were synthesized from luteolin (3',4',5,7-tetrahydroxyflavone) and fatty acid chlorides. These acylated semi-biomolecules show a hypsochromic shift in UV-Vis spectra of about Δλ(A→B)=58nm and absorbed in the centre of the harmful UV-B band (λ(max)=295nm). Luteolin was also hydroxyethylated with Br(CH(2))(2)OH. This substitution has no effect on the λ(max)=330nm absorption of luteolin (UV-A band). Finally the natural 4'-O-β-glucosyl-3',5,7-trihydroxyflavone was extracted from edelweiss and used as a purified natural benchmark. Glycosylated and hydroxyethylated luteolin are both UV stable. Fully acylated luteolin derivatives degrade upon UV exposure to a stable UV-C screen with a hypsochroic shift Δλ(B→C)=35nm. All in all, three molecular structures based on luteolin with sunscreen properties were found, distinguishable in: UV-A, UV-B, and UV-C filters. The natural product based UV-absorbers show promise as alternatives to synthetic molecules and nanoparticles in sunscreen products.     

The role of UV-B radiation in aquatic and terrestrial ecosystems--an experimental and functional analysis of the evolution of UV-absorbing compounds

We analysed and compared the functioning of UV-B screening pigments in plants from marine, fresh water and terrestrial ecosystems, along the evolutionary line of cyanobacteria, unicellular algae, primitive multicellular algae, charophycean algae, lichens, mosses and higher plants, including amphibious macrophytes. Lichens were also included in the study. We were interested in the following key aspects: (a) does the water column function effectively as an 'external UV-B filter'?; (b) do aquatic plants need less 'internal UV-B screening' than terrestrial plants?; (c) what role does UV screening play in protecting the various plant groups from UV-B damage, such as the formation of thymine dimers?; and (d) since early land 'plants' (such as the predecessors of present-day cyanobacteria, lichens and mosses) experienced higher UV-B fluxes than higher plants, which evolved later, are primitive aquatic and land organisms (cyanobacteria, algae, lichens, mosses) better adapted to present-day levels of UV-B than higher plants? Furthermore, polychromatic action spectra for the induction of UV screening pigments of aquatic organisms have been determined. This is relevant for translating 'physical' radiation measurements of solar UV-B into 'biological' and 'ecological' effects. From the action spectra, radiation amplification factors (RAFs) have been calculated. These action spectra allow us to determine any mitigating or antagonistic effects in the ecosystems and therefore qualify the damage prediction for the ecosystems under study. We summarize and discuss the main results based on three years of research of four European research groups. The central theme of the work was the investigation of the effectiveness of the various screening compounds from the different species studied in order to gain some perspective of the evolutionary adaptations from lower to higher plant forms. The induction of mycosporine-like amino acids (MAAs) was studied in the marine dinoflagellate Gyrodinium dorsum, the green algal species Prasiola stipitata and in the cyanobacterium Anabaena sp. While visible (400-700 nm) and long wavelength UV-A (315-400 nm) showed only a slight effect, MAAs were effectively induced by UV-B (280-315 nm). The growth of the lower land organisms studied, i.e. the lichens Cladina portentosa, Cladina foliacaea and Cladonia arbuscula, and the club moss Lycopodiumannotinum, was not significantly reduced when grown under elevated UV-B radiation (simulating 15% ozone depletion). The growth in length of the moss Tortula ruralis was reduced under elevated UV-B. Of the aquatic plants investigated the charophytes Chara aspera showed decreased longitudinal growth under elevated UV-B. In the 'aquatic higher plants' studied, Ceratophyllum demersum, Batrachium trichophyllum and Potamogeton alpinus, there was no such depressed growth with enhanced UV-B. In Chara aspera, neither MAAs nor flavonoids could be detected. Of the terrestrial higher plants studied, Fagopyrum esculentum, Deschampsia antarctica, Vicia faba, Calamagrostis epigejos and Carex arenaria, the growth of the first species was depressed with enhanced UV-B, in the second species length growth was decreased, but the shoot number was increased, and in the latter two species of a dune grassland there was no reduced growth with enhanced UV-B. In the dune grassland species studied outdoors, at least five different flavonoids appeared in shoot tissue. Some of the flavonoids in the monocot species, which were identified and quantified with HPLC, included orientin, luteolin, tricin and apigenin. A greenhouse study with Vicia faba showed that two flavonoids (aglycones) respond particularly to enhanced UV-B. Of these, quercetin is UV-B inducible and mainly located in epidermal cells, while kaempferol occurs constitutively. In addition to its UV-screening function, quercetin may also act as an antioxidant. Polychromatic action spectra were determined for induction of the UV-absorbing pigments in three photosynthetic organisms, representing very different taxonomic groups and different habitats. In ultraviolet photobiology, action spectra mainly serve two purposes: (1) identification of the molecular species involved in light absorption; and (2) calculation of radiation amplification factors for assessing the effect of ozone depletion. Radiation amplification factors (RAFs) were calculated from the action spectra. In a somewhat simplified way, RAF can be defined as the percent increase of radiation damage for a 1% depletion of the ozone layer. Central European summer conditions were used in the calculations, but it has been shown that RAF values are not critically dependent on latitude or season. If only the ultraviolet spectral region is considered, the RAF values obtained are 0.7 for the green alga Prasiola stipitata, 0.4 for the dinoflagellate Gyrodinium dorsum, and 1.0 for the cyanobacterium Anabaena sp. In the case of P. stipitata, however, the effect of visible light (PAR, photosynthetically active radiation, 400-700 nm) is sufficient to lower the RAF to about 0.4, while the PAR effect for G. dorsum is negligible. RAFs for some damage processes, such as for DNA damage (RAF=2.1 if protective effects or photorepair are not considered [1]), are higher than those above. Our interpretation of this is that if the ozone layer is depleted, increased damaging radiation could overrule increased synthesis of protective pigments. In addition to investigating the functional effectiveness of the different screening compounds, direct UV effects on a number of key processes were also studied in order to gain further insight into the ability of the organisms to withstand enhanced UV-B radiation. To this end, the temperature-dependent repair of cyclobutane dimers (CPD) and (6-4) photoproducts induced by enhanced UV-B was studied in Nicotiana tabacum, and the UV-B induction of CPD was studied in the lichen Cladonia arbuscula. Also, photosynthesis and motility were monitored and the response related to the potential function of the screening compounds of the specific organism.     

Mycosporine-like amino acids in the marine dinoflagellate Gyrodinium dorsum: induction by ultraviolet irradiation

Cultures of the marine dinoflagellate Gyrodinium dorsum have been exposed to polychromatic radiation (photosynthetically active radiation and UV) from a solar simulator for up to 72 h. Different irradiance spectra in the ultraviolet are produced by inserting cut-off filters between lamp and samples. The mycosporine-like amino acid (MAA) content and composition are investigated by spectroscopic and chromatographic analysis. The study reveals that G. dorsum contains a complex mixture of several aminocyclohexenimine-MAAs and one aminocyclohexenone-MAA. UV irradiation around 320 nm induces an increase in the concentration of all MAAs in the samples. In contrast, exposure to short-wavelength UV-B radiation results in decreased overall MAA production. Furthermore, there is a spectral shift in the absorption of the MAA mixture towards shorter wavelengths, indicating that short-wavelength UV-B induces an altered MAA composition. The amount of MAAs is normalized to the chlorophyll a concentration.     

Phototaxis in the cyanobacterium Synechocystis sp. PCC 6803: role of different photoreceptors

The second cyanobacterial phytochrome Cph2 from Synechocystis sp. PCC 6803 was suggested as a part of a light-stimulated signal transduction chain inhibiting movement toward blue light. Cph2 has the two bilin binding sites, cysteine-129 and cysteine-1022, that might be involved in sensing of red/far-red and blue light, respectively. Here, we present data on wavelength dependence of the phototaxis inhibition under blue light, indicating that Cph2 itself is the photoreceptor for this blue light response. We found that inhibition of blue-light phototaxis in wild-type cells occurred below the transition point of about 470 nm. Substitution of cysteine-1022 with valine led to photomovement of the cells toward blue light (cph2(-) mutant phenotype). Analysis of mutants lacking cysteine-129 in the N-terminal chromophore binding domain indicated that this domain is also important for Cph2 function or folding of the protein. Furthermore, putative blue-light and phytochrome-like photoreceptors encoded by the Synechocystis sp. PCC 6803 genome were inactivated in wild-type and cph2 knockout mutant background. Our results suggest that none of these potential photoreceptors interfere with Cph2 function, although inactivation of taxD1 as well as slr1694 encoding a BLUF protein led to cells that reversed the direction of movement under blue light illumination in mutant strains of cph2.     

Stereochemistry of the reduction step mediated by recombinant 1-deoxy-D-xylulose 5-phosphate isomeroreductase

[formula: see text] The stereochemistry of the 1-deoxy-D-xylulose 5-phosphate (DXP) isomeroreductase reduction step has been examined using the recombinant enzyme from Synechocystis sp. PCC6803. Using [3-2H]DXP and [4S-2H]NADPH, it has been determined that the C1 pro-S hydrogen in the 2-C-methyl-D-erythritol 4-phosphate product derives from C3 of DXP, indicating that hydride attack occurs on the re face of the intermediate aldehyde. The 4S-hydride from NADPH is delivered, assigning this enzyme as a class B dehydrogenase.     

A phytochrome-like protein AphC triggers the cAMP signaling induced by far-red light in the cyanobacterium Anabaena sp. strain PCC7120

In the filamentous, nitrogen-fixing cyanobacterium Anabaena sp. PCC7120, red light (630 nm) decreased, whereas far-red light (720 nm) increased cellular adenosine 3',5'-cyclic monophosphate (cAMP) content. To find a red and far-red light photoreceptor that triggers the cAMP signal cascade, we disrupted 10 open reading frame having putative chromophore-binding GAF domains. The response of the cellular cAMP concentration to red and far-red light in each open reading frame disruptant was determined. It was found that only the mutant of the gene all2699 failed to respond to far-red light. The open reading frame named as aphC encoded a protein with 920 amino acids including GAF domains similar to those involved in Cph2, a photoreceptor of Synechocystis sp. PCC6803. To determine which adenylate cyclase (AC) is responsible for far-red light signal, we disrupted all AC genes and found that CyaC was the candidate. The enzymatic activity of CyaC might be controlled by a far-red light photoreceptor through the phosphotransfer reaction. The site-specific mutant of the Asp59 residue of the receiver (R1) domain of CyaC lost its light-response capability. It was suggested that the far-red light signal was received by AphC and then transferred to the N-terminal response regulator domain of CyaC. Then its catalytic activity was stimulated, which increased the cellular cAMP concentration and drove the subsequent signal transduction cascade.