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In this study, a cyanobacteria-bacteria consortium containing native wastewater bacteria and immobilized Synechococcus sp. was constructed. The cyanobacterial cellular responses (including growth, biomass and lipid productivity) and contaminant removal ability (for TN, TP, COD and antibiotics) in the consortium were evaluated during the advanced treatment of wastewater containing 10-50 μg/L of mixed antibiotics (amoxicillin, tetracycline, erythromycin, sulfadiazine and ciprofloxacin) with the addition of a certain phytohormone (indole-3-acetic acid, gibberellin A3 or 6-benzylaminopurine) at trace level within a period of four days. Each phytohormone promoted the growth of Synechococcus sp. and increased the tolerance of Synechococcus sp. to mixed antibiotics. Indole-3-acetic acid coupled to moderate antibiotic stress could elevate lipid productivity and lipid content of Synechococcus sp. to 33.50 mg/L/day and 43.75%, respectively. Phytohormones increased the pollutant removal performance of the cyanobacteria-bacteria consortium through the stimulation of cyanobacterial growth and the regulation of cyanobacteria-bacteria interaction, which increased the abundances of microalgae-associated bacteria including Flavobacterium, Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, Bosea, Sphingomonas and Emticicia. Up to 80.83%, 98.06%, 83.26%, 99.84%, 99.50%, 89.41%, 65.61% and 60.65% of TN, TP, COD, amoxicillin, tetracycline, erythromycin, sulfadiazine and ciprofloxacin were removed by the consortium with the addition of phytohormones. In general, indole-3-acetic acid was the optimal phytohormone for enhancing lipid production and contaminant removal performance of the cyanobacteria-bacteria consortium., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Cyanobacteria are among the most suitable organisms for the capture of excessive amounts of CO 2 and can be grown in extreme environments. In our research we use the single-celled freshwater cyanobacteria Synechococcus elongatus PCC7942 PAMCOD strain and Synechocystis sp. PCC6714 for the production of carbohydrates and hydrogen. PAMCOD strain and Synechocystis sp. PCC6714 synthesize sucrose when exposed to salinity stress, as their main compatible osmolyte. We examined the cell proliferation rate and the sucrose accumulation in those two different strains of cyanobacteria under salt (0.4 M NaCl) and heat stress (35 0 C) conditions. The intracellular sucrose (mol sucrose content per Chl a) was found to increase by 50% and 108% in PAMCOD strain and Synechocystis sp. PCC6714 cells, respectively. As previously reported, PAMCOD strain has the ability to produce hydrogen through the process of dark anaerobic fermentation (Vayenos D, Romanos GE, Papageorgiou GC, Stamatakis K (2020) Photosynth Res 146, 235-245). In the present study, we demonstrate that Synechocystis sp. PCC6714 has also this ability. We further examined the optimal conditions during the dark fermentation of PAMCOD and Synechocystis sp. PCC6714 regarding H 2 formation, increasing the PAMCOD H 2 productivity from 2 nmol H 2 h - 1 mol Chl a - 1 to 23 nmol H 2 h - 1 mol Chl a - 1 . Moreover, after the dark fermentation, the cells demonstrated proliferation in both double BG-11 and BG-11 medium enriched in NaNO 3 , thus showing the sustainability of the procedure., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Microcystin-LR (MC-LR) is a harmful cyanotoxin that inhibits 1 and 2A serine-threonine protein phosphatases. This study examines the influence of MC-LR on chloroplast division and the underlying mechanisms and consequences in Arabidopsis. MC-LR increased the frequency of dividing chloroplasts in hypocotyls in a time range of 1-96 h. At short-term exposures to MC-LR, small-sized chloroplasts (longitudinal diameters ≤6 μm) were more sensitive to these stimulatory effects, while both small and large chloroplasts showed stimulations at long-term exposure. After 48 h, the cyanotoxin increased the frequency of small-sized chloroplasts, indicating the stimulation of division. MC-LR inhibited protein phosphatases in whole hypocotyls and isolated chloroplasts, while it did not induce oxidative stress. We show for the first time that total cellular phosphatases play important roles in chloroplast division and that particular chloroplast phosphatases may be involved in these processes. Interestingly, MC-LR has a protective effect on cyanobacterial division during methyl-viologen (MV) treatments in Synechococcus PCC6301. MC-LR production has harmful effects on ecosystems and it may have an ancient cell division regulatory role in stressed cyanobacterial cells, the evolutionary ancestors of chloroplasts. We propose that cytoplasmic (eukaryotic) factors also contribute to the relevant effects of MC-LR in plants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Heat shock protein 90 (Hsp90) is essential for eukaryotic cells, whereas bacterial homologs play a role under stresses and in pathogenesis. Identifying species-specific Hsp90 inhibitors is challenging because Hsp90 is evolutionarily conserved. We found that a cyclic lipopeptide surfactin inhibits the ATPase activity of Hsp90 from the cyanobacterium Synechococcus elongatus (S.elongatus) PCC 7942 but does not inhibit Escherichia coli (E.coli), yeast and human Hsp90s. Molecular docking simulations indicated that surfactin could bind to the N-terminal dimerization interface of the cyanobacterial Hsp90 in the ATP- and ADP-bound states, which provided molecular insights into the species-selective inhibition. The data suggest that surfactin inhibits a rate-limiting conformational change of S.elongatus Hsp90 in the ATP hydrolysis. Surfactin also inhibited the interaction of the cyanobacterial Hsp90 with a model substrate, and suppressed S.elongatus growth under heat stress, but not that of E.coli. Surfactin did not show significant cellular toxicity towards mammalian cells. These results indicate that surfactin inhibits the cellular function of Hsp90 specifically in the cyanobacterium. The present study shows that a cyclic peptide has a great specificity to interact with a specific homolog of a highly conserved protein family., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.)

In cyanobacteria, the PII protein (the glnB gene product) regulates a number of proteins involved in nitrogen assimilation including PipX, the coactivator of the global nitrogen regulator protein NtcA. In Synechococcus elongatus PCC 7942, construction of a PII-less mutant retaining the wild-type pipX gene is difficult because of the toxicity of uncontrolled action of PipX and the other defect(s) resulting from the loss of PIIper se, but the nature of the PipX toxicity and the PipX-independent defect(s) remains unclear. Characterization of a PipX-less glnB mutant (PD4) in this study showed that the loss of PII increases the sensitivity of PSII to ammonium. Ammonium was shown to stimulate the formation of reactive oxygen species in the mutant cells. The ammonium-sensitive growth phenotype of PD4 was rescued by the addition of an antioxidant α-tocopherol, confirming that photo-oxidative damage was the major cause of the growth defect. A targeted PII mutant retaining wild-type pipX was successfully constructed from the wild-type S. elongatus strain (SPc) in the presence of α-tocopherol. The resulting mutant (PD1X) showed an unusual chlorophyll fluorescence profile, indicating extremely slow reduction and re-oxidation of QA, which was not observed in mutants defective in both glnB and pipX. These results showed that the aberrant action of uncontrolled PipX resulted in an impairment of the electron transport reactions in both the reducing and oxidizing sides of QA., (© The Author(s) 2021. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Characterizing the cell-level metabolic trade-offs that phytoplankton exhibit in response to changing environmental conditions is important for predicting the impact of these changes on marine food web dynamics and biogeochemical cycling. The time-selective proteome-labeling approach, bioorthogonal noncanonical amino acid tagging (BONCAT), has potential to provide insight into differential allocation of resources at the cellular level, especially when coupled with proteomics. However, the application of this technique in marine phytoplankton remains limited. We demonstrate that the marine cyanobacteria Synechococcus sp. and two groups of eukaryotic algae take up the modified amino acid l-homopropargylglycine (HPG), suggesting that BONCAT can be used to detect translationally active phytoplankton. However, the impact of HPG addition on growth dynamics varied between groups of phytoplankton. In addition, proteomic analysis of Synechococcus cells grown with HPG revealed a physiological shift in nitrogen metabolism, general protein stress, and energy production, indicating a potential limitation for the use of BONCAT in understanding the cell-level response of Synechococcus sp. to environmental change. Variability in HPG sensitivity between algal groups and the impact of HPG on Synechococcus physiology indicates that particular considerations should be taken when applying this technique to other marine taxa or mixed marine microbial communities. IMPORTANCE Phytoplankton form the base of the marine food web and substantially impact global energy and nutrient flow. Marine picocyanobacteria of the genus Synechococcus comprise a large portion of phytoplankton biomass in the ocean and therefore are important model organisms. The technical challenges of environmental proteomics in mixed microbial communities have limited our ability to detect the cell-level adaptations of phytoplankton communities to a changing environment. The proteome labeling technique, bioorthogonal noncanonical amino acid tagging (BONCAT), has potential to address some of these challenges by simplifying proteomic analyses. This study explores the ability of marine phytoplankton to take up the modified amino acid, l-homopropargylglycine (HPG), required for BONCAT, and investigates the proteomic response of Synechococcus to HPG. We not only demonstrate that cyanobacteria can take up HPG but also highlight the physiological impact of HPG on Synechococcus , which has implications for future applications of this technique in the marine environment.

Marine plastic pollution is a growing concern worldwide and has the potential to impact marine life via leaching of chemicals, with zinc (Zn), a common plastic additive, observed at particularly high levels in plastic leachates in previous studies. At this time, however, little is known regarding how elevated Zn affects key groups of marine primary producers. Marine cyanobacterial genera Prochlorococcus and Synechococcus are considered to be some of the most abundant oxygenic phototrophs on earth, and together contribute significantly to oceanic primary productivity. Here we set out to investigate how two Prochlorococcus (MIT9312 and NATL2A) and two Synechococcus (CC9311 and WH8102) strains, representative of diverse ecological niches, respond to exposure to high Zn concentrations. The two genera showed differences in the timing and degree of growth and physiological responses to elevated Zn levels, with Prochlorococcus strains showing declines in their growth rate and photophysiology following exposure to 27 µg l -1 Zn, while Synechococcus CC9311 and WH8102 growth rates declined significantly on exposure to 52 and 152 µg l -1 Zn, respectively. Differences were also observed in each strain's capacity to maintain cell wall integrity on exposure to different levels of Zn. Our results indicate that excess Zn has the potential to pose a challenge to some marine picocyanobacteria and highlights the need to better understand how different marine Prochlorococcus and Synechococcus strains may respond to increasing concentrations of Zn in some marine regions.

Cyanobacteria hold promise as cell factories for the photoautotrophic conversion of carbon dioxide to useful chemicals. For the eventual commercial viability of such processes, cyanobacteria need to be engineered for (i) efficient channeling of carbon flux toward the product of interest and (ii) improved product tolerance, the latter being the focus of this study. We chose the recently reported, fast-growing, high light and CO 2 tolerant cyanobacterium Synechococcus elongatus PCC 11801 for adaptive laboratory evolution. In two parallel experiments that lasted over 8400 h of culturing and 100 serial passages, S. elongatus PCC 11801 was evolved to tolerate 5 g/L n-butanol or 30 g/L 2,3-butanediol representing a 100% improvement in concentrations tolerated. The evolved strains retained alcohol tolerance even after being passaged several times without the alcohol stress suggesting that the changes were permanent. Whole genome sequencing of the n-butanol evolved strains revealed mutations in a number of stress responsive genes encoding translation initiation factors, RpoB and an ABC transporter. In 2,3-butanediol evolved strains, genes for ClpC, a different ABC transporter, glyceraldehyde-3-phosphate dehydrogenase and phosphoribulokinase were found to be mutated. Furthermore, the evolved strains showed significant improvement in tolerance toward several other alcohols. Notably, the n-butanol evolved strain could tolerate up to 32 g/L ethanol, thereby making it a promising host for photosynthetic production of biofuels via metabolic engineering., (Crown Copyright © 2020. Published by Elsevier B.V. All rights reserved.)

The aim and novelty of this paper are found in assessing the influence of inhibitors and antibiotics on intact cell MALDI-TOF mass spectra of the cyanobacterium Synechococcus sp. UPOC S4 and to check the impact on reliability of identification. Defining the limits of this method is important for its use in biology and applied science. The compounds included inhibitors of respiration, glycolysis, citrate cycle, and proteosynthesis. They were used at 1-10 μM concentrations and different periods of up to 3 weeks. Cells were also grown without inhibitors in a microgravity because of expected strong effects. Mass spectra were evaluated using controls and interpreted in terms of differential peaks and their assignment to protein sequences by mass. Antibiotics, azide, and bromopyruvate had the greatest impact. The spectral patterns were markedly altered after a prolonged incubation at higher concentrations, which precluded identification in the database of reference spectra. The incubation in microgravity showed a similar effect. These differences were evident in dendrograms constructed from the spectral data. Enzyme inhibitors affected the spectra to a smaller extent. This study shows that only a long-term presence of antibiotics and strong metabolic inhibitors in the medium at 10 -5 M concentrations hinders the correct identification of cyanobacteria by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF).

Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942 exhibit dissimilar tolerance to Cr(VI) with a tenfold difference in their EC 50 value for Cr(VI). This contrasting tolerance was attributed to the difference in the ability to transport Cr(VI) and to detoxify ROS. The present study used biochemical assays and chlorophyll fluorescence to investigate the effect of growth with Cr(VI) on photosynthesis in the two cyanobacteria. In absence of Cr(VI), all the measured parameters viz ., rates of CO 2 fixation, PSII and PSI activities were higher in Synechocystis in comparison to Synechococcus , suggesting intrinsic differences in their photosynthesis. Growth in the presence of Cr(VI) reduced the pigment content and photosystems' activities in both cyanobacteria. It was further observed that photosynthetic functions were more adversely affected in Synechocystis in comparison to Synechococcus, in spite of exposure to tenfold lower Cr(VI) concentration. The effective quantumyield of PSII and PSI obtained by chlorophyll fluorescence measurements increased in the presence of Cr(VI) in Synechococcus whereas it decreased in Synechocystis . However, the overall CO 2 fixation remained unchanged. These results indicated that, in addition to the intrinsic difference in photosynthetic rates, the two cyanobacteria exhibit differential modulation of photosynthetic machinery upon Cr(VI) exposure and Synechococcus could adapt better it's photosystems to counter the oxidative stress.

In several marine hosts of microalgae, fluorescent natural products may play an important role. While the ecological function of these compounds is not well understood, an interaction of these molecules with the photosynthesis of the symbionts has been suggested. In this study, the effect of Ageladine A (Ag A), a pH-dependent fluorophore found in sponges of the genus Agelas, on microalgal fluorescence was examined. The spectra showed an accumulation of Ag A within the cells, but with variable impacts on fluorescence. While in two Synechococcus strains, fluorescence of phycoerythrin increased significantly, the fluorescence of other Synechococcus strains was not affected. In four out of the five eukaryote species examined, chlorophyll a (Chl a) fluorescence intensity was modulated. In Tisochrysis lutea, for example, the position of the fluorescence emission maximum of Chl a was shifted. The variety of these effects of Ag A on microalgal fluorescence suggests that fluorophores derived from animals could play a crucial role in shaping the composition of marine host/symbiont systems., Competing Interests: The authors have declared that no competing interests exist.

Motivation: Determining intracellular metabolic flux through isotope labeling techniques such as 13C metabolic flux analysis (13C-MFA) incurs significant cost and effort. Previous studies have shown transcriptomic data coupled with constraint-based metabolic modeling can determine intracellular fluxes that correlate highly with 13C-MFA measured fluxes and can achieve higher accuracy than constraint-based metabolic modeling alone. These studies, however, used validation data limited to E. coli and S. cerevisiae grown on glucose, with significantly similar flux distribution for central metabolism. It is unclear whether those results apply to more diverse metabolisms, and therefore further, extensive validation is needed., Results: In this paper, we formed a dataset of transcriptomic data coupled with corresponding 13C-MFA flux data for 21 experimental conditions in different unicellular organisms grown on varying carbon substrates and conditions. Three computational flux-balance analysis (FBA) methods were comparatively assessed. The results show when uptake rates of carbon sources and key metabolites are known, transcriptomic data provides no significant advantage over constraint-based metabolic modeling (average correlation coefficients, transcriptomic E-Flux2 0.725 and SPOT 0.650 vs non-transcriptomic pFBA 0.768). When uptake rates are unknown, however, predictions obtained utilizing transcriptomic data are generally good and significantly better than those obtained using constraint-based metabolic modeling alone (E-Flux2 0.385 and SPOT 0.583 vs pFBA 0.237). Thus, transcriptomic data coupled with constraint-based metabolic modeling is a promising method to obtain intracellular flux estimates in microorganisms, particularly in cases where uptake rates of key metabolites cannot be easily determined, such as for growth in complex media or in vivo conditions., Competing Interests: The authors have declared that no competing interests exist.

Monitoring of oxidative stress caused by a wide range of reactive oxygen species (ROS) is essential to have an idea about the fitness and growth of photosynthetic organisms. The imaging-based oxidative stress measurement in cyanobacteria using 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) dye has the limitation of small sample size as the only selected number of cells are analyzed to measure the ROS levels. Here, we developed a method for oxidative stress measurement by DCFH-DA and flow cytometer (FCM) using unicellular Synechococcus elongatus PCC 7942 and filamentous Fremyella diplosiphon BK14 cyanobacteria. F. diplosiphon BK14 inherently possess high levels of ROS and showed higher sensitivity to hydrogen peroxide treatment in comparison to S. elongatus PCC 7942. We successfully measured oxidative stress in glutaredoxin lacking strain (Δgrx3) of S. elongatus PCC 7942, and wild-type Synechocystis sp. PCC 6803 using FCM based method. Importantly, ROS were not detected in these two strains of cyanobacteria by fluorescence microscope-based method due to their small spherical morphology. Δgrx3 strain showed high ROS levels in comparison to its wild-type strain. Treatment of abiotic factors such as high PAR in wild-type and Δgrx3 strains of S. elongatus PCC 7942, low PAR or low PAR + UVR in wild-type S. elongatus PCC 7942, and high PAR or high PAR + NaCl in Synechocystis sp. PCC 6803 increased oxidative stress. In summary, the FCM based method can measure ROS levels produced due to physiological conditions associated with genetic changes or abiotic stress in a large population of cells regardless of their morphology. Therefore, the present study shows the usefulness of the method in monitoring the health of organisms in a large scale cultivation system., Competing Interests: Declaration of competing interest All authors declare no conflict of interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Microplastics or plastic particles less than 5 mm in size are a ubiquitous and damaging pollutant in the marine environment. However, the interactions between these plastic particles and marine microorganisms are just starting to be understood. The objective of this study was to measure the responses of a characteristic marine organism (Synechococcus sp. PCC 7002) to an anthropogenic stressor (polyethelene nanoparticles and microparticles) using molecular techniques. This investigation showed that polyethylene microparticles and nanoparticles have genetic, enzymatic and morphological effects on Synechococcus sp. PCC 7002. An RT-PCR analysis showed increases in the expression of esterase and hydrolase genes at 5 days of exposure to polyethylene nanoparticles and at 10 days of exposure to polyethylene microparticles. A qualitative enzymatic assay also showed esterase activity in nanoparticle exposed samples. Cryo-scanning electron microscopy was used to assess morphological changes in exopolymer formation resulting from exposure to polyethylene microparticles and nanoparticles. The data from this paper suggests that microplastic and nanoplastics could be key microbial stressors and should be investigated in further detail., Competing Interests: The authors have declared that no competing interests exist.

Mathematical models can enable a predictive understanding of mechanism in cell biology by quantitatively describing complex networks of interactions, but such models are often poorly constrained by available data. Owing to its relative biochemical simplicity, the core circadian oscillator in Synechococcus elongatus has become a prototypical system for studying how collective dynamics emerge from molecular interactions. The oscillator consists of only three proteins, KaiA, KaiB, and KaiC, and near-24-h cycles of KaiC phosphorylation can be reconstituted in vitro. Here, we formulate a molecularly detailed but mechanistically naive model of the KaiA-KaiC subsystem and fit it directly to experimental data within a Bayesian parameter estimation framework. Analysis of the fits consistently reveals an ultrasensitive response for KaiC phosphorylation as a function of KaiA concentration, which we confirm experimentally. This ultrasensitivity primarily results from the differential affinity of KaiA for competing nucleotide-bound states of KaiC. We argue that the ultrasensitive stimulus-response relation likely plays an important role in metabolic compensation by suppressing premature phosphorylation at nighttime., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

1-Butanol production using photosynthetic organisms such as cyanobacteria has garnered interest among researchers due to its high potential as a sustainable biofuel. Previously, the cyanobacterium Synechococcus elongatus PCC 7942 was engineered to produce 1-butanol through the introduction of a modified CoA-dependent pathway. S. elongatus strain DC11, a high producer of 1-butanol, was constructed based on metabolomics-assisted strain engineering. DC11 can reach a production titer of 418.7 mg/L in 6 days, cutting the production time in half compared to the previously constructed DC7. Regardless, the final 1-butanol titer of DC11 was still low compared to other microbial hosts. Sensitivity towards 1-butanol of the producing strain has been known as one of main hurdles for improving cyanobacterial production system. Thus, to improve cyanobacterial-based 1-butanol production in the future, we employed the metabolomics approach to study the intrinsic effect of improved 1-butanol productivity in DC11. This study focused on metabolite profiling of DC11 using LC/MS/MS. Results showed that there is an accumulation of disaccharide-P and sucrose/trehalose in DC11 compared to the DC7. These metabolites were previously reported to have a role in salt and alcohol stress response in cyanobacteria and therefore, DC11 was subjected to 0.2 M of NaCl and 1000 mg/L of 1-butanol for further investigation. DC11 with stress treatment showed a more prominent accumulation of sugars and nucleosides compared to control. The results obtained from this study may be beneficial for future strain improvement strategies in S. elongatus, particularly addressing the metabolic response of this strain upon 1-butanol stress., (Copyright © 2019 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

Recent synthetic biology efforts have raised biosafety concerns for possible release of engineered cyanobacteria into natural environments. To address the issues, we developed a controllable metal ion induced biocontainment system for two model cyanobacteria. First, six ion-inducible promoters were respectively evaluated in both Synechococcus elongatus PCC 7942 and the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973, leading to the identification of an iron ion-repressed promoter P isiAB with low leakage and a reduction-fold of 5.4 and 7.9, respectively. Second, holin-endolysin and nuclease NucA systems were introduced, the inhibition rate of which against two Synechococcus strains varied from 61% to 86.4%. Third, two toxin/antitoxin modules were identified capable of inducing programmed suicide in both Synechococcus strains after induction. Furthermore, an escape experiment was conducted and the results showed that the system was able to achieve an escape frequency below the detection limit of 10 -9 after 3 days' duration, demonstrating the strategy integrating iron ion-inducible promoter P isiAB and that toxin/antitoxin modules could be a useful tool for cyanobacterium biocontainment.

Ethylene is a volatile alkene which is used in large commercial scale as a precursor in plastic industry, and is currently derived from petroleum refinement. As an alternative production strategy, photoautotrophic cyanobacteria Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942 have been previously evaluated as potential biotechnological hosts for producing ethylene directly from CO 2 , by the over-expression of ethylene forming enzyme (efe) from Pseudomonas syringae. This work addresses various open questions related to the use of Synechococcus as the engineering target, and demonstrates long-term ethylene production at rates reaching 140 µL L -1  h -1 OD 750 -1 without loss of host vitality or capacity to produce ethylene. The results imply that the genetic instability observed earlier may be associated with the expression strategies, rather than efe over-expression, ethylene toxicity or the depletion of 2-oxoglutarate-derived cellular precursors in Synechococcus. In context with literature, this study underlines the critical differences in expression system design in the alternative hosts, and confirms Synechococcus as a suitable parallel host for further engineering.

Mycosporine-like amino acids (MAAs) are a class of well-documented UV-screening compounds produced by taxonomically diverse organisms. Extensive studies revealed that a rare MAA, mycosporine-2-glycine (M2G), possesses unique biological activities and functions. M2G is not only a potent antioxidant, but also suppresses protein glycation in vitro, and production of inflammatory mediators in RAW 264.7 macrophages. The present study evaluates vital functions of M2G in a heterologous expression system. The stress-sensitive fresh water cyanobacterium Synechococcus elongatus PCC7942, carrying a M2G biosynthetic gene cluster, was generated. The M2G-expressing cells were more tolerant to H2O2-induced oxidative stress than the wild type, with a half-maximal inhibitory concentration (IC50) value of 2.3 ± 0.06 mM. Transcriptional analysis revealed that all M2G biosynthetic genes were highly up-regulated under oxidative stress. Further, expression of vital genes in the cellular antioxidant defense system, including sodB, cat and tpxA were modulated and up-regulated. Elevated M2G was detected under oxidative stress as well as salt stress treatments. This study provides insight into the molecular and cellular effects of the M2G biosynthetic gene cluster, contributing to understanding of the mechanism behind physiological plasticity under this heterologous expression system., (© FEMS 2019.)

The environmental considerations attributing to the escalation of carbon dioxide emissions have raised alarmingly. Consequently, the concept of sequestration and biological conversion of CO 2 by photosynthetic microorganisms is gaining enormous recognition. In this study, in an attempt to discern the synergistic CO 2 tolerance mechanisms, metabolic responses to increasing CO 2 concentrations were determined for Synechococcus elongatus PCC 11801, a fast-growing, novel freshwater strain, using quantitative proteomics. The protein expression data revealed that the organism responded to elevated CO 2 by not only regulating the cellular transporters involved in carbon-nitrogen uptake and assimilation but also by inducing photosynthesis, carbon fixation and glycolysis. Several components of photosynthetic machinery like photosystem reaction centers, phycobilisomes, cytochromes, etc. showed a marked up-regulation with a concomitant downshift in proteins involved in photoprotection and redox maintenance. Additionally, enzymes belonging to the TCA cycle and oxidative pentose phosphate pathway exhibited a decline in their expression, further highlighting that the demand for reduced cofactors was fulfilled primarily through photosynthesis. The present study brings the first-ever comprehensive assessment of intricate molecular changes in this novel strain while shifting from carbon-limited to carbon-sufficient conditions and may pave the path for future host and pathway engineering for production of sustainable fuels through efficient CO 2 capture.

Pico-cyanobacteria and micro-cyanobacteria coexist ubiquitously in many lakes. Differences in cell size and abilities to utilize nutrients may influence their distribution patterns. In this study, Synechococcus sp. and Microcystis aeruginosa were chosen as pico- and micro-cyanobacteria, respectively. Gradient phosphorus treatments (0.002, 0.01, 0.05, and 0.25 mg P L -1 ) were designed in mono- and co-cultures. Growth curves were recorded and fitted by the Monod equation. Moreover, the interspecific competition was analyzed by the Lotka-Volterra model. When mono-cultured in lower P conditions (≤ 0.01 mg P L -1 ), Synechococcus sp. obtained much higher biomass than M. aeruginosa. But, M. aeruginosa grew faster than Synechococcus sp. in higher P groups (≥ 0.05 mg P L -1 ) (p < 0.05). Synechococcus sp. has abilities to thrive in low-phosphorus environments, whereas M. aeruginosa favored high-phosphorus conditions. In co-cultures, Synechococcus sp. strongly inhibited M. aeruginosa at each P treatment.

The light harvesting photosystem in cyanobacteria offers a potential pathway for the regeneration of the nicotinamide cofactor NADPH, thereby facilitating the application of cyanobacteria as excellent whole cell biocatalysts in oxidoreductase-mediated biotransformation. The use of cyanobacterial metabolism for cofactor recycling improves the atom economy of the process compared to the commonly employed enzyme-coupled cofactor recycling using enzymes such as glucose dehydrogenase. Here we report the asymmetric conversion of acetophenone to chiral 1-phenylethanol by recombinant Synechococcus elongatus PCC 7942 whole cell biocatalyst that expresses the NADPH dependent L. kefir alcohol dehydrogenase. Besides light, it was observed that carbon dioxide levels play a critical role in improving the bioconversion efficiency possibly due to the enhanced growth rate and improved cofactor availability at elevated CO 2 levels. Complete reduction of acetophenone to optically pure (R)-1-phenylethanol at 99% enantiomeric excess was achieved within 6 h with a relatively low cell density of 0.66 g/l by coupling optimum light and CO 2 levels and without the need for a co-substrate., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The role of macroalgal allelopathy in aquatic systems has received increasing attention as a potential means of controlling cyanobacterial blooms. However, the allelopathic activity of Chara sp. on coexisting and bloom-forming picocyanobacteria is still largely unknown. Therefore, the laboratory experiments were conducted to investigate the allelopathic activity of extracts of Chara aspera, C. baltica, and C. canescens on the growth, the fluorescence parameters: maximum and effective quantum yield of photosystem II (PSII) photochemistry (F v /F m and ΦPSII, respectively) and photosynthesis parameters such as the initial slope of photosynthesis-irradiance (P-E) curves (alpha) and photosynthetic capacity (P m ) of the picocyanobacterium Synechococcus sp. Batch cultures of picocyanobacterium were exposed to three concentrations of extracts originating from three charophyte cultures and the effect was followed at three sampling times. Dried specimens of C. aspera, C. baltica, and C. canescens were extracted in the water-based matrix and the initial Synechococcus sp. inoculum, derived from unialgal culture media, was used. We found both negative and positive allelopathic effects of all tested Chara extracts on Synechococcus sp. The strongest adverse impact of picocyanobacterium growth was caused by C. baltica. This study clearly demonstrated that the allelopathic effect depends on the Chara species identity. Our results also suggested that some allelopathic Chara sp. have the potential to mitigate harmful cyanobacterial blooms in systems dominated by Synechococcus sp.

Cyanobacteria are photosynthetic prokaryotes that have been extensively studied as potential autotrophic biotechnological hosts for the production of different carbon-based end-products directly from atmospheric CO 2 . While commercially competitive applications do not yet exist, the production of ethanol in cyanobacteria is the most mature technology, endorsed by relatively high production yields and established status of ethanol in the global biofuel market. Within this concept, the aim here was to systematically compare ethanol tolerance of different commonly used cyanobacterial strains and substrains, in order to assess their relative potential for biotechnological production platforms. The comparison revealed clear strain-specific differences in ethanol toxicity, with growth inhibition GI 50 values ranging between 3 g L -1 (0.4% V/V) and 28 g L -1 (3.5% V/V). The most tolerant wild-type strains were Synechocystis sp. PCC 6803 (substrain A) and Synechococcus sp. PCC 7002, which did not show any apparent effect in growth below ethanol concentrations 9.2 g L -1 (1.2% V/V). In comparison to typical biotechnological yeast strains used for ethanol fermentation, these values are clearly lower but still around the same order of magnitude. The results also underlined the challenges in direct number-based comparison between cyanobacterial strains and culture conditions due to inconsistencies in respect to chlorophyll content, cell morphology and optical properties., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Culture experiments were conducted on ten phytoplankton species to examine their biological and physiological responses during exposure to oil and a combination of oil and dispersant. The species tested included a range of taxa typically found in the Gulf of Mexico such as cyanobacteria, chlorophytes, and diatoms. Cultures were exposed to Macondo surrogate oil using the water accommodated fraction (WAF), and dispersed oil using a chemically enhanced WAF (CEWAF) and diluted CEWAF, to replicate conditions following the Deepwater Horizon spill in the Gulf of Mexico. A range of responses were observed, that could broadly class the algae as either "robust" or "sensitive" to oil and/or dispersant exposure. Robust algae were identified as Synechococcus elongatus, Dunaliella tertiolecta, two pennate diatoms Phaeodactylum tricornutum and Navicula sp., and Skeletonema grethae CCMP775, and were largely unaffected by any of the treatments (no changes to growth rate or time spent in lag phase relative to controls). The rest of the phytoplankton, all centric diatoms, exhibited at least some combination of reduced growth rates or increased lag time in response to oil and/or dispersant exposure. Photophysiology did not have a strong treatment effect, with significant inhibition of photosynthetic efficiency (F v /F m ) only observed in the CEWAF, if at all. We found that the effects of oil and dispersants on phytoplankton physiology were species-dependent, and not always detrimental. This has significant implications on how oil spills might impact phytoplankton community structure and bloom dynamics in the Gulf of Mexico, which in turn impacts higher trophic levels., (© 2018 Phycological Society of America.)

Nitrogenase is an oxygen-vulnerable metalloenzyme that catalyzes nitrogen fixation. It largely remains unknown how nitrogenase coexists with oxygenic photosynthesis in nonheterocystous cyanobacteria, since there have been no appropriate model cyanobacteria so far. Here, we demonstrate in vivo transposon tagging in the nonheterocystous cyanobacterium Leptolyngbya boryana as a forward genetics approach. By conjugative transfer, a mini-Tn5-derived vector, pKUT-Tn5-Sm/Sp, was transferred from Escherichia coli to L. boryana cells. Of 1839 streptomycin-resistant colonies, we isolated three mutants showing aberrant diazotrophic growth. Genome resequencing identified the insertion sites of the transposon in the mutants. This in vivo transposon tagging mutagenesis of L. boryana provides a promising system to investigate molecular mechanisms to resolve the Oxygen Paradox between nitrogen fixation and oxygenic photosynthesis in cyanobacteria., (© 2018 Federation of European Biochemical Societies.)

Compared to extensive studies reporting the aluminum (Al) toxicity to terrestrial plants and freshwater organisms, very little is known about how marine phytoplankton responds to Al in the field. Here we report the marine phytoplankton responses to Al enrichment in the South China Sea (SCS) using on-deck bottle incubation experiments during eight cruises from May 2010 to November 2013. Generally, Al addition alone enhanced the growth of diatom and Trichodesmium, and nitrogen fixation, but it inhibited the growth of dinoflagellates and Synechococcus. Nevertheless, Al addition alone did not influence the chlorophyll a concentration of the entire phytoplankton assemblages. By adding nitrate and phosphate simultaneously, Al enrichment led to substantial increases in chlorophyll a concentration (especially that of the picophytoplankton<3μm), and cell abundances of diatom and photosynthetic picoeukaryotes. These results indicate varied responses of phytoplankton in different size fractions and taxonomic groups to Al enrichment. Further, by simultaneously adding different macronutrients and/or sufficient trace metals including iron, we found that the phytoplankton responses to Al enrichment were relevant to nutrients coexisting in the environment. Al enrichment may give some phytoplankton a competitive edge over using nutrients, especially the limited ones. The possible influences of Al on the competitors and grazers (predators) of some phytoplankton might indirectly contribute to the positive responses of the phytoplankton to Al enrichment. Our results indicate that Al may influence marine carbon cycle by impacting phytoplankton growth and structure in natural seawater., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Sucrose and glycogen syntheses in cyanobacteria share the common precursor glucose-1-phosphate. It is generally assumed that lowering glycogen synthesis could drive more carbon toward sucrose synthesis that can be induced by salt stress among cyanobacteria. By using a theophylline-dependent riboswitch system, the expression of glgC , a key gene in glycogen synthesis, was downregulated in a quantitative manner in a sucrose-secreting strain of Synechococcus elongatus PCC 7942. We observed that the stepwise suppression of glycogen synthesis limited rather than stimulated sucrose production in the salt-stressed cells, suggesting that glycogen could serve as a carbon pool for the synthesis of sucrose. Accordingly, we generated glycogen-overproducing strains, but the increased glycogen pool alone did not stimulate sucrose production, indicating that alternative steps limit the carbon flux toward the synthesis of sucrose. Consistent with previous studies that showed that sucrose-phosphate synthase (SPS) catalyzes the rate-limiting step in sucrose synthesis, the combination of glycogen overproduction and sps overexpression resulted in increased sucrose production. Our results indicate that the glycogen and sucrose pools are closely linked in Synechococcus elongatus PCC 7942, and we propose that enhancing the glycogen pool could be a promising strategy for the improvement of sucrose production by cyanobacteria in the presence of a strong sucrose synthesis sink. IMPORTANCE Many cyanobacteria naturally synthesize and accumulate sucrose when stressed by NaCl, which provides novel possibilities for obtaining sugar feedstock by engineering of cyanobacteria. It has been assumed that glycogen synthesis competes with sucrose synthesis for the carbon flux. However, our results showed that the suppression of glycogen synthesis decreased rather than stimulated sucrose production in a sucrose-secreting strain of Synechococcus elongatus PCC 7942. This result suggests that glycogen could serve as a supportive rather than a competitive carbon pool for the synthesis of sucrose, providing new insights about the relation between glycogen synthesis and sucrose synthesis in cyanobacteria. This finding is also useful to guide metabolic engineering work to optimize the production of sucrose and possibly other products by cyanobacteria., (Copyright © 2018 American Society for Microbiology.)

Background: The photosynthetic apparatus is targeted by various herbicides, including several amides such as diuron and linuron. Considering the need for the discovery of new active ingredients to cope with weed resistance, the synthesis of a series of trifluoromethyl aryl amides is herein described whose inhibitory properties were assessed in vitro on the photosynthetic electron transport chain, and in vivo on the growth of a model cyanobacterial strain. Theoretical studies were also carried out., Results: Starting with 1-fluoro-2-nitro-4-(trifluoromethyl) benzene, the preparation of the amides was achieved via a three-step sequence, namely nucleophilic aromatic substitution, reduction with SnCl 2 /HCl, and acylation reactions. The measurement of ferricyanide reduction by functionally intact spinach chloroplasts showed that several derivatives are capable of inhibiting the photosynthetic apparatus. The most active amides presented IC 50 values close to 1 μmol L -1 , and showed the presence of a 4-bromophenyl group as a common structural feature. The addition of these brominated amides to the culture medium of a model cyanobacterial strain, Synechococcus elongatus PCC 6301, caused various degrees of growth inhibition. Theoretical studies (molecular modeling and quantitative structure-activity relationship) of all amides and their comparison with some known herbicides confirmed these experimental findings and provided more in-depth information about the possible molecular target of these compounds., Conclusion: Trifluoromethyl amides herein described, which were shown to act at the PSII level, may represent a novel scaffold to be exploited aiming at the development of new active ingredients for weed control. © 2017 Society of Chemical Industry., (© 2017 Society of Chemical Industry.)

Enhancement of alcohol tolerance in microorganisms is an important strategy for improving bioalcohol productivity. Although cyanobacteria can be used as a promising biocatalyst to produce various alcohols directly from CO 2 , low productivity, and low tolerance against alcohols are the main issues to be resolved. Nevertheless, to date, a mutant with increasing alcohol tolerance has rarely been reported. In this study, we attempted to select isopropanol (IPA)-tolerant mutants of Synechococcus elongatus PCC 7942 using UV-C-induced random mutagenesis, followed by enrichment of the tolerant candidates in medium containing 10 g/L IPA and screening of the cells with a high growth rate in the single cell culture system in liquid medium containing 10 g/L IPA. We successfully acquired the most tolerant strain, SY1043, which maintains the ability to grow in medium containing 30 g/L IPA. The photosynthetic oxygen-evolving activities of SY1043 were almost same in cells after 72 h incubation under light with or without 10 g/L IPA, while the activity of the wild-type was remarkably decreased after the incubation with IPA. SY1043 also showed higher tolerance to ethanol, 1-butanol, isobutanol, and 1-pentanol than the wild type. These results suggest that SY1043 would be a promising candidate to improve alcohol production using cyanobacteria. Biotechnol. Bioeng. 2017;114: 1771-1778. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Phytoplankton constitute an important component of surface water ecosystems; however little is known about their contribution to biotransformation of organic micropollutants. To elucidate biotransformation processes, batch experiments with two cyanobacterial species (Microcystis aeruginosa and Synechococcus sp.) and one green algal species (Chlamydomonas reinhardtii) were conducted. Twenty-four micropollutants were studied, including 15 fungicides and 9 pharmaceuticals. Online solid phase extraction (SPE) coupled with liquid chromatography (LC)-high resolution tandem mass spectrometry (HRMS/MS) was used together with suspect and nontarget screening to identify transformation products (TPs). 14 TPs were identified for 9 micropollutants, formed by cytochrome P450-mediated oxidation, conjugation and methylation reactions. The observed transformation pathways included reactions likely mediated by promiscuous enzymes, such as glutamate conjugation to mefenamic acid and pterin conjugation of sulfamethoxazole. For 15 compounds, including all azole fungicides tested, no TPs were identified. Environmentally relevant concentrations of chemical stressors had no influence on the transformation types and rates.

The ability of cyanobacteria to survive many environmental stress factors is a testament to their resilience in nature. Of these environmental stress factors, overexposure to zinc is important to study since excessive zinc intake can be a severe hazard. Zinc toxicity in freshwater has been demonstrated to affects organisms such as invertebrates, algae and cyanobacteria. Cyanobacteria which possess increased resistance to zinc have been isolated. It is therefore important to elucidate the mechanism of survival and response to determine what factors allow their survival; as well as any remediation implications they may have. To characterize the effects of zinc in freshwater cyanobacteria, we investigated the response of Synechococcus sp. IU 625 (S. IU 625) over 29days to various concentrations (10, 25, and 50mg/L) of ZnCl 2 . S. IU 625 was shown to be tolerant up to 25mg/L ZnCl 2 exposure, with 10mg/L ZnCl 2 having no outward physiological change and 50mg/L ZnCl 2 proving lethal to the cells. To determine a potential mechanism Inductive Coupled Plasma-Mass Spectrometry (ICP-MS) and RNA-seq analysis were performed on zinc exposed cells. Analysis performed on days 4 and 7 indicated that response is dose-dependent, with 10mg/L ZnCl 2 exhibiting nearly all zinc extracellular, corresponding with upregulation of cation transport response. Whereas the 25mg/L ZnCl 2 exhibited half of total zinc sequestered by the cells, which corresponds with the upregulation of sequestering proteins such as metallothionein and the downregulation of genes involved with ATP synthesis and phycobilisome assembly. These analyses were combined with growth monitoring, microscopy, quantitative polymerase chain reaction (qPCR) and flow cytometry to present a full spectrum of mechanisms behind zinc response in S. IU 625., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Exopolymer particles are found throughout the ocean and play a significant biogeochemical role in carbon cycling. Transparent exopolymer particles (TEP) are composed of acid polysaccharides, and Coomassie staining particles (CSP) are proteins. TEPs have been extensively studied in the ocean, while CSP have been largely overlooked. The objective of this research was to determine the role of stress and cell permeability in the formation of TEP and CSP. The diatom Thalassiosira weissflogii and cyanobacterium Synechococcus elongatus were grown in batch cultures and exposed to hydrogen peroxide (0, 10, and 100 μM) as an environmental stressor. There was no correlation between TEP and CSP concentrations, indicating that they are different populations of particles rather than different chemical components of the same particles. CSP concentrations were not affected by hydrogen peroxide concentration and did not correlate with indicators of stress and cell death. In contrast, TEP concentrations in both taxa were correlated with a decrease in the effective quantum yield of photosystem II, increased activity of caspase-like enzymes, and an increase in the proportion of the population with permeable cell membranes, indicating that TEP production was associated with the process of cell death. These data show that different environmental factors and physiological processes affected the production of TEP and CSP by phytoplankton. TEP and CSP are separate populations of exopolymer particles with potentially different biogeochemical roles in the ocean., (© 2016 Phycological Society of America.)

The assimilation of N-NO 3 - requires more energy than that of N-NH 4 + . This becomes relevant when energy is limiting and may impinge differently on cell energy budget depending on depth, time of the day and season. We hypothesize that N-limited and energy-limited cells of the oceanic cyanobacterium Synechococcus sp. differ in their response to the N source with respect to growth, elemental stoichiometry and carbon allocation. Under N limitation, cells retained almost absolute homeostasis of elemental and organic composition, and the use of NH 4 + did not stimulate growth. When energy was limiting, however, Synechococcus grew faster in NH 4 + than in NO 3 - and had higher C (20%), N (38%) and S (30%) cell quotas. Furthermore, more C was allocated to protein, whereas the carbohydrate and lipid pool size did not change appreciably. Energy limitation also led to a higher photosynthetic rate relative to N limitation. We interpret these results as an indication that, under energy limitation, the use of the least expensive N source allowed a spillover of the energy saved from N assimilation to the assimilation of other nutrients. The change in elemental stoichiometry influenced C allocation, inducing an increase in cell protein, which resulted in a stimulation of photosynthesis and growth., (© 2016 John Wiley & Sons Ltd.)

Picocyanobacteria from the genus Synechococcus are ubiquitous in ocean waters. Their phylogenetic and genomic diversity suggests ecological niche differentiation, but the selective forces influencing this are not well defined. Marine picocyanobacteria are sensitive to Cu toxicity, so adaptations to this stress could represent a selective force within, and between, 'species', also known as clades. Here, we compared Cu stress responses in cultures and natural populations of marine Synechococcus from two co-occurring major mesotrophic clades (I and IV). Using custom microarrays and proteomics to characterize expression responses to Cu in the lab and field, we found evidence for a general stress regulon in marine Synechococcus. However, the two clades also exhibited distinct responses to copper. The Clade I representative induced expression of genomic island genes in cultures and Southern California Bight populations, while the Clade IV representative downregulated Fe-limitation proteins. Copper incubation experiments suggest that Clade IV populations may harbour stress-tolerant subgroups, and thus fitness tradeoffs may govern Cu-tolerant strain distributions. This work demonstrates that Synechococcus has distinct adaptive strategies to deal with Cu toxicity at both the clade and subclade level, implying that metal toxicity and stress response adaptations represent an important selective force for influencing diversity within marine Synechococcus populations., (© 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Singlet-excited oxygen ( 1 O 2 * ) has been recognized as the most destructive member of the reactive oxygen species (ROS) which are formed during oxygenic photosynthesis by plants, algae, and cyanobacteria. ROS and 1 O 2 * are known to damage protein and phospholipid structures and to impair photosynthetic electron transport and de novo protein synthesis. Partial protection is afforded to photosynthetic organism by the β-carotene (β-Car) molecules which accompany chlorophyll (Chl) a in the pigment-protein complexes of Photosystem II (PS II). In this paper, we studied the effects of exogenously added β-Car on the initial kinetic rise of Chl a fluorescence (10-1000 μs, the OJ segment) from the unicellular cyanobacterium Synechococcus sp. PCC7942. We show that the added β-Car enhances Chl a fluorescence when it is excited at an intensity of 3000 μmol photons m -2 s -1 but not when excited at 1000 μmol photons m -2 s -1 . Since β-Car is an efficient scavenger of 1 O 2 * , as well as a quencher of 3 Chl a * (precursor of 1 O 2 * ), both of which are more abundant at higher excitations, we assume that the higher Chl a fluorescence in its presence signifies a protective effect against photo-oxidative damages of Chl proteins. The protective effect of added β-Car is not observed in O 2 -depleted cell suspensions. Lastly, in contrast to β-Car, a water-insoluble molecule, a water-soluble scavenger of 1 O 2 * , histidine, provides no protection to Chl proteins during the same time period (10-1000 μs).

Cyanobacterial mutants defective in acyl-acyl carrier protein synthetase (Aas) secrete free fatty acids (FFAs) into the external medium and hence have been used for the studies aimed at photosynthetic production of biofuels. While the wild-type strain of Synechocystis sp. PCC 6803 is highly sensitive to exogenously added linolenic acid, mutants defective in the aas gene are known to be resistant to the externally provided fatty acid. In this study, the wild-type Synechocystis cells were shown to be sensitive to lauric, oleic, and linoleic acids as well, and the resistance to these fatty acids was shown to be enhanced by inactivation of the aas gene. On the basis of these observations, we developed an efficient method to isolate aas-deficient mutants from cultures of Synechocystis cells by counter selection using linoleic acid or linolenic acid as the selective agent. A variety of aas mutations were found in about 70 % of the FFA-resistant mutants thus selected. Various aas mutants were isolated also from Synechococcus sp. PCC 7002, using lauric acid as a selective agent. Selection using FFAs was useful also for construction of markerless aas knockout mutants from Synechocystis sp. PCC 6803 and Synechococcus sp. PCC 7002. Thus, genetic engineering of FFA-producing cyanobacterial strains would be greatly facilitated by the use of the FFAs for counter selection., Competing Interests: The authors declare that they have no conflict of interest. Ethical approval This article does not contain any studies with human participants or animals performed by any of the authors.

The photoinhibition of photosystem I (PSI) is lethal to oxygenic phototrophs. Nevertheless, it is unclear how photodamage occurs or how oxygenic phototrophs prevent it. Here, we provide evidence that keeping P700 (the reaction center chlorophyll in PSI) oxidized protects PSI. Previous studies have suggested that PSI photoinhibition does not occur in the two model cyanobacteria, Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942, when photosynthetic CO 2 fixation was suppressed under low CO 2 partial pressure even in mutants deficient in flavodiiron protein (FLV), which mediates alternative electron flow. The lack of FLV in Synechococcus sp. PCC 7002 (S. 7002), however, is linked directly to reduced growth and PSI photodamage under CO 2 -limiting conditions. Unlike Synechocystis sp. PCC 6803 and S. elongatus PCC 7942, S. 7002 reduced P700 during CO 2 -limited illumination in the absence of FLV, resulting in decreases in both PSI and photosynthetic activities. Even at normal air CO 2 concentration, the growth of S. 7002 mutant was retarded relative to that of the wild type. Therefore, P700 oxidation is essential for protecting PSI against photoinhibition. Here, we present various strategies to alleviate PSI photoinhibition in cyanobacteria., (© 2016 American Society of Plant Biologists. All Rights Reserved.)

The residues of antineoplastic drugs are considered as new and emerging pollutants in aquatic environments. Recent experiments showed relatively high toxicity of 5-fluorouracil (5-FU), imatinib mesylate (IM), etoposide (ET) and cisplatin (CP) that are currently among most widely used antineoplastic drugs, against phytoplankton species. In this study, we investigated the toxic potential of the mixture of 5-FU + IM + ET against green alga Pseudokirchneriella subcapitata and cyanobacterium Synechococcus leopoliensis, and the stability and sorption of these drugs to algal cells. Toxic potential of the mixture was predicted by the concepts of 'concentration addition' and 'independent action' and compared to the experimentally determined toxicity. In both test species, the measured toxicity of the mixture was at effects concentrations EC10-EC50 higher than the predicted, whereas at higher effect concentration (EC90), it was lower. In general, P. subcapitata was more sensitive than S. leopoliensis. The stability studies of the tested drugs during the experiment showed that 5-FU, IM and CP are relatively stable, whereas in the cultures exposed to ET, two transformation products with the same mass as ET but different retention time were detected. The measurements of the cell-linked concentrations of the tested compounds after 72 h exposure indicated that except for CP (1.9 % of the initial concentration), these drugs are not adsorbed or absorbed by algal cells. The results of this study showed that in alga and cyanobacteria exposure to the mixture of 5-FU + ET + IM, in particular at low effect concentration range, caused additive or synergistic effect on growth inhibition, and they suggest that single compound toxicity data are not sufficient for the proper toxicity prediction for aquatic primary producers.

Accumulation of electrons under conditions of environmental stress produces a reduced state in the photosynthetic electron transport (PET) system and causes the reduction of O 2 by PSI in the thylakoid membranes to produce the reactive oxygen species superoxide radical, which irreversibly inactivates PSI. This study aims to elucidate the molecular mechanism for the oxidation of reaction center Chl of PSI, P700, after saturated pulse (SP) light illumination of the cyanobacterium Synechococcus elongatus PCC 7942 under steady-state photosynthetic conditions. Both P700 and NADPH were transiently oxidized after SP light illumination under CO 2 -depleted photosynthesis conditions. In contrast, the Chl fluorescence intensity transiently increased. Compared with the wild type, the increase in Chl fluorescence and the oxidations of P700 and NADPH were greatly enhanced in a mutant (Δflv1/3) deficient in the genes encoding FLAVODIIRON 1 (FLV1) and FLV3 proteins even under high photosynthetic conditions. Furthermore, oxidation of Cyt f was also observed in the mutant. After SP light illumination, a transient suppression of O 2 evolution was also observed in Δflv1/3. From these observations, we propose that the reduction in the plastquinone (PQ) pool suppresses linear electron flow at the Cyt b 6 /f complex, which we call the reduction-induced suppression of electron flow (RISE) in the PET system. The accumulation of the reduced form of PQ probably suppresses turnover of the Q cycle in the Cyt b 6 /f complex., (© The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Sensitivity of four tropical cyanobacteria viz. Coelosphaerium sp., Synechococcus sp., Oscillatoria sp. and Chroococcus sp. to environmentally relevant concentrations of Cr(6+), Cd(2+) and Zn(2+)was assessed based on fluorescence change as a proxy for growth reduction. At 24 h exposure, the growth reduction inthe cyanobacteria followed the order: Zn(2+) < Cr(6+) ≤ Cd(2+). Of the four cyanobacteria, Synechococcus was the most sensitive for Cr(6+), where as Chroococcus was the most sensitive for Cd(2+)and Zn(2+). Sensitivity was gradually decreased by 96 h implying the acquisition of tolerance by cyanobacteria to heavy metal ions with prolonged exposure.

Cyanobacteria have evolved effective adaptive mechanisms to improve photosynthesis and CO2 fixation. The central CO2-fixing machinery is the carboxysome, which is composed of an icosahedral proteinaceous shell encapsulating the key carbon fixation enzyme, Rubisco, in the interior. Controlled biosynthesis and ordered organization of carboxysomes are vital to the CO2-fixing activity of cyanobacterial cells. However, little is known about how carboxysome biosynthesis and spatial positioning are physiologically regulated to adjust to dynamic changes in the environment. Here, we used fluorescence tagging and live-cell confocal fluorescence imaging to explore the biosynthesis and subcellular localization of β-carboxysomes within a model cyanobacterium, Synechococcus elongatus PCC7942, in response to light variation. We demonstrated that β-carboxysome biosynthesis is accelerated in response to increasing light intensity, thereby enhancing the carbon fixation activity of the cell. Inhibition of photosynthetic electron flow impairs the accumulation of carboxysomes, indicating a close coordination between β-carboxysome biogenesis and photosynthetic electron transport. Likewise, the spatial organization of carboxysomes in the cell correlates with the redox state of photosynthetic electron transport chain. This study provides essential knowledge for us to modulate the β-carboxysome biosynthesis and function in cyanobacteria. In translational terms, the knowledge is instrumental for design and synthetic engineering of functional carboxysomes into higher plants to improve photosynthesis performance and CO2 fixation., (© 2016 American Society of Plant Biologists. All Rights Reserved.)

Marine environments are frequently exposed to oil spills as a result of transportation, oil drilling or fuel usage. Whereas large oil spills and their effects have been widely documented, more common and recurrent small spills typically escape attention. To fill this important gap in the assessment of oil-spill effects, we performed two independent supervised full sea releases of 5 m(3) of crude oil, complemented by on-board mesocosm studies and sampling of accidentally encountered slicks. Using rapid on-board biological assays, we detect high bioavailability and toxicity of dissolved and dispersed oil within 24 h after the spills, occurring fairly deep (8 m) below the slicks. Selective decline of marine plankton is observed, equally relevant for early stages of larger spills. Our results demonstrate that, contrary to common thinking, even small spills have immediate adverse biological effects and their recurrent nature is likely to affect marine ecosystem functioning.

The chlorophyll a fluorescence based antimicrobial susceptibility testing (AST) method presented in a previous work was based on the measurement of Chl a fluorescence of the gram(-) cyanobacterium Synechococcus sp. PCC 7942. Synechococcus sp. PCC 7942 as a gram(-) bacterium is affected by antibacterial agents via mechanisms affecting all gram(-) bacteria, however, as an exclusively phototrophic organism it would also be affected by photosynthesis inhibitory action of an agent that otherwise has no antibacterial properties. In this report, the method is modified by replacing the exclusively phototrophic Synechococcus sp. PCC 7942 with the Synechocystis sp. PCC 6714, capable of both phototrophic and heterotrophic growth in order to add versatility and better reflect the antibacterial effects of surfaces under study towards nonphotosynthetic bacteria., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Cyanobacteria can be utilized as a potential biocatalyst for the production of biofuels and biochemicals directly from CO2. Useful mutants of cyanobacteria, which can grow rapidly or are resistant to specific metabolic products, are essential to improve the productivity of biofuels. In this study, we developed a single cell culture system to effectively screen mutant cyanobacteria using magnetite nanoparticles and magnetic force. Lens culinaris Agglutinin (LCA) was selected as a lectin, which binds to the surface of Synechococcus elongatus PCC7942 cells and the LCA-conjugated magnetite cationic liposomes (MCLs) were developed for magnetic labeling of PCC7942 cells. The MCL-labeled PCC7942 cells were magnetically patterned at a single cell level by using 6,400 iron pillars of the pin-holder device. The device enabled 1,600 single cells to be arrayed in one square centimeter. We cultured the patterned cells in liquid medium and achieved higher colony-forming ratio (78.4%) than that obtained using conventional solid culture method (4.8%). Single cells with different properties could be distinguished in the single cell culture system depending on their growth. Furthermore, we could selectively pick up the target cells and subsequently perform efficient isolation culture. The ratio of successful isolation culture using the developed method was 13 times higher than that of the conventional methods. Thus, the developed system would serve as a powerful tool for screening mutant cyanobacteria., (© 2015 Wiley Periodicals, Inc.)

Marine Synechococcus are some of the most diverse and ubiquitous phytoplankton, and iron (Fe) is an essential micronutrient that limits productivity in many parts of the ocean. To investigate how coastal and oceanic Atlantic Synechococcus strains acclimate to Fe availability, we compared the growth, photophysiology, and quantitative proteomics of two Synechococcus strains from different Fe regimes. Synechococcus strain WH8102, from a region in the southern Sargasso Sea that receives substantial dust deposition, showed impaired growth and photophysiology as Fe declined, yet used few acclimation responses. Coastal WH8020, from the dynamic, seasonally variable New England shelf, displayed a multitiered, hierarchical cascade of acclimation responses with different Fe thresholds. The multitiered response included changes in Fe acquisition, storage, and photosynthetic proteins, substitution of flavodoxin for ferredoxin, and modified photophysiology, all while maintaining remarkably stable growth rates over a range of Fe concentrations. Modulation of two distinct ferric uptake regulator (Fur) proteins that coincided with the multitiered proteome response was found, implying the coastal strain has different regulatory threshold responses to low Fe availability. Low nitrogen (N) and phosphorus (P) availability in the open ocean may favor the loss of Fe response genes when Fe availability is consistent over time, whereas these genes are retained in dynamic environments where Fe availability fluctuates and N and P are more abundant.

Excess sulfite is well known to have toxic effects on photosynthetic activities and growth in plants, however, so far, the behavior of the photosynthetic apparatus during sulfite-stress has not been characterized as to the responsible proteins or genes. Here, the effects of sulfite on photosystem complexes were investigated in a cyanobacterium, Synechococcus elongatus PCC 7942, a possible model organism of chloroplasts. Culturing of the cells for 24 h in the presence of 10 mM sulfite retarded cell growth of the wild type, concomitantly with synthesis of Chl and phycobilisome repressed. The excess sulfite simultaneously repressed photosynthesis by more than 90%, owing largely to structural destabilization and resultant inactivation of the PSII complex, which seemed to consequently retard the cell growth. Notably, the PsbO protein, one of the subunits that construct the water-splitting system of PSII, was retained at a considerable level, and disruption of the psbO gene led to higher sensitivity of photosynthesis and growth to sulfite. Meanwhile, the PSI complex showed monomerization of its trimeric configuration with little effect on the activity. The structural alterations of these PS complexes depended on light. Our data provide evidence for quantitative decreases in the photosystem complex(es) including their antenna(e), structural alterations of the PSI and PSII complexes that would modulate their functions, and a crucial role of psbO in PSII protection, in Synechococcus cells during sulfite-stress. We suggest that the reconstruction of the photosystem complexes is beneficial to cell survival., (© The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Currently, cyanobacteria are regarded as potential biofuel sources. Large-scale cultivation of cyanobacteria in seawater is of particular interest because seawater is a low-cost medium. In the present study, we examined differences in light-harvesting and energy transfer processes in the cyanobacterium Synechococcus sp. PCC 7002 grown in different cultivation media, namely modified A medium (the optimal growth medium for Synechococcus sp. PCC 7002) and f/2 (a seawater medium). The concentrations of nitrate and phosphate ions were varied in both media. Higher nitrate ion and/or phosphate ion concentrations yielded high relative content of phycobilisome. The cultivation medium influenced the energy transfers within phycobilisome, from phycobilisome to photosystems, within photosystem II, and from photosystem II to photosystem I. We suggest that the medium also affects charge recombination at the photosystem II reaction center and formation of a chlorophyll-containing complex.

Two unicellular cyanobacteria, Synechocystis and Synechococcus, showed contrasting tolerance to Cr(VI); with Synechococcus being 12-fold more tolerant than Synechocystis to potassium dichromate. The mechanism responsible for this differential sensitivity to Cr(VI) was explored in this study. Total content of photosynthetic pigments as well as photosynthetic activity decreased at lower concentration of Cr(VI) in Synechocystis as compared to Synechococcus. Experiments with (51)Cr showed Cr to accumulate intracellularly in both the cyanobacteria. At lower concentrations, Cr(VI) caused excessive ROS generation in Synechocystis as compared to that observed in Synechococcus. Intrinsic levels of enzymatic antioxidants, i.e., superoxide dismutase, catalase and 2-Cys-peroxiredoxin were considerably higher in Synechococcus than Synechocystis. Content of total thiols (both protein as well as non-protein) and reduced glutathione (GSH) was also higher in Synechococcus as compared to Synechocystis. This correlated well with higher content of carbonylated proteins observed in Synechocystis than Synechococcus. Additionally, in contrast to Synechocystis, Synechococcus exhibited better tolerance to other oxidative stresses like high intensity light and H2O2. The data indicate that the disparity in the ability to detoxify ROS could be the primary mechanism responsible for the differential tolerance of these cyanobacteria to Cr(VI)., (Copyright © 2015 Elsevier B.V. All rights reserved.)

In this work, a recombinant cyanobacterium strain with increased photosynthesis rate, cell growth and carbohydrate production efficiency was genetically engineered by co-expressing ictB, ecaA, and acsAB (encoded for bacterial cellulose) in Synechococcus elongatus PCC7942. The resulting cyanobacterial biomass could be effectively hydrolyzed with dilute acid (2% sulfuric acid), achieving a nearly 90% glucose recovery at a biomass concentration of 80 g/L. Bioethanol can be produced from fermenting the acidic hydrolysate of S. elongatus PCC7942 via separate hydrolysis and fermentation (SHF) process at a concentration of 7.2 g/L and with a 91% theoretical yield., (Copyright © 2014 Elsevier Ltd. All rights reserved.)