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Concept: Synechocystis

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Bacterial phototaxis was first recognized over a century ago, but the method by which such small cells can sense the direction of illumination has remained puzzling. The unicellular cyanobacterium Synechocystis sp. PCC 6803 moves with Type IV pili and measures light intensity and color with a range of photoreceptors. Here, we show that individual Synechocystis cells do not respond to a spatiotemporal gradient in light intensity, but rather they directly and accurately sense the position of a light source. We show that directional light sensing is possible because Synechocystis cells act as spherical microlenses, allowing the cell to see a light source and move towards it. A high-resolution image of the light source is focused on the edge of the cell opposite to the source, triggering movement away from the focused spot. Spherical cyanobacteria are probably the world’s smallest and oldest example of a camera eye.

Concepts: DNA, Cyanobacteria, Archaea, Bacteria, Organism, Light, Synechocystis, Pilus

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Abstract In the present economy, difficulties to access energy sources are real drawbacks to maintain our current lifestyle. In fact, increasing interests have been gathered around efficient strategies to use energy sources that do not generate high CO2 titers. Thus, science-funding agencies have invested more resources into research on hydrogen among other biofuels as interesting energy vectors. This article reviews present energy challenges and frames it into the present fuel usage landscape. Different strategies for hydrogen production are explained and evaluated. Focus is on biological hydrogen production; fermentation and photon-fuelled hydrogen production are compared. Mathematical models in biology can be used to assess, explore and design production strategies for industrially relevant metabolites, such as biofuels. We assess the diverse construction and uses of genome-scale metabolic models of cyanobacterium Synechocystis sp. PCC6803 to efficiently obtain biofuels. This organism has been studied as a potential photon-fuelled production platform for its ability to grow from carbon dioxide, water and photons, on simple culture media. Finally, we review studies that propose production strategies to weigh this organism’s viability as a biofuel production platform. Overall, the work presented in this review unveils the industrial capabilities of cyanobacterium Synechocystis sp. PCC6803 to evolve interesting metabolites as a clean biofuel production platform.

Concepts: Oxygen, Carbon dioxide, Metabolism, Hydrogen, Yeast, Synechocystis, Hydrogen production, Biological hydrogen production

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BACKGROUND: The transcriptomes of several cyanobacterial strains have been shown to exhibit diurnal oscillation patterns, reflecting the diurnal phototrophic lifestyle of the organisms.The analysis of such genome-wide transcriptional oscillations is often facilitated by the use of clustering algorithms in conjunction with a number of pre-processing steps. Biological interpretation is usually focussed on the time and phase of expression of the resulting groups of genes.However, the use of microarray technology in such studies requires the normalization of pre-processing data, with unclear impact on the qualitative and quantitative features of the derived information on the number of oscillating transcripts and their respective phases. RESULTS: A microarray based evaluation of diurnal expression in the cyanobacterium Synechocystis sp. PCC 6803 is presented. As expected, the temporal expression patterns reveal strong oscillations in transcript abundance.We compare the Fourier transformation-based expression phase before and after the application of quantile normalization, median polishing, cyclical LOESS, and least oscillating set (LOS) normalization.Whereas LOS normalization mostly preserves the phases of the raw data, the remaining methods introduce systematic biases. In particular, quantile-normalization is found to introduce a phase-shift of 180°, effectively changing night-expressed genes into day-expressed ones. Comparison of a large number of clustering results of differently normalized data shows that the normalization method determines the result. Subsequent steps, such as the choice of data transformation, similarity measure, and clustering algorithm, only play minor roles.We find that the standardization and the DTF transformation are favorable for the clustering of time series in contrast to the 12m transformation. We use the cluster-wise functional enrichment of a clustering derived by LOS normalization, clustering using flowClust, and DFT transformation to derive the diurnal biological program of Synechocystis sp.. CONCLUSION: Application of quantile normalization, median polishing, and also cyclic LOESS normalization of the presented cyanobacterial dataset lead to increased numbers of oscillating genes and the systematic shift of the expression phase. The LOS normalization minimizes the observed detrimental effects. As previous analyses employed a variety of different normalization methods, a direct comparison of results must be treated with caution.

Concepts: Cyanobacteria, Gene, Gene expression, Transcription, Molecular biology, Oscillation, Synechocystis, Seasonality

1

Environmental cues can stimulate a variety of single-cell responses, as well as collective behaviors that emerge within a bacterial community. These responses require signal integration and transduction, which can occur on a variety of time scales and often involve feedback between processes, for example, between growth and motility. Here, we investigate the dynamics of responses of the phototactic, unicellular cyanobacterium Synechocystis sp. PCC6803 to complex light inputs that simulate the natural environments that cells typically encounter. We quantified single-cell motility characteristics in response to light of different wavelengths and intensities. We found that red and green light primarily affected motility bias rather than speed, while blue light inhibited motility altogether. When light signals were simultaneously presented from different directions, cells exhibited phototaxis along the vector sum of the light directions, indicating that cells can sense and combine multiple signals into an integrated motility response. Under a combination of antagonistic light signal regimes (phototaxis-promoting green light and phototaxis-inhibiting blue light), the ensuing bias was continuously tuned by competition between the wavelengths, and the community response was dependent on both bias and cell growth. The phototactic dynamics upon a rapid light shift revealed a wavelength dependence on the time scales of photoreceptor activation/deactivation. Thus, Synechocystis cells achieve exquisite integration of light inputs at the cellular scale through continuous tuning of motility, and the pattern of collective behavior depends on single-cell motility and population growth.IMPORTANCE The photosynthetic cyanobacterium Synechocystis sp. exhibits phototaxis that is dependent on the incident light wavelength through the action of various photoreceptors. In natural environments, cells experience a set of highly dynamic and complex light inputs, yet how cells transduce multiple or dynamic inputs into motion is unknown. In this study, we measured the phototactic behaviors of single cells and communities as a function of light intensity or when illuminated by combinations of lights of different wavelengths or incidence directions. Responses to a spectrum of light regimes revealed that Synechocystis sp. integrates information about the light environment to tune its phototactic response, which is likely generated by competition among photoreceptors and the degree of wavelength-regulated growth to sensitively control the direction and degree of movement.

Concepts: Cyanobacteria, Photosynthesis, Bacteria, Light, Electromagnetic radiation, Wavelength, Synechocystis, Visible spectrum

1

Bacterial toxin-antitoxin (TA) systems are genetic elements, which are encoded by plasmid as well as chromosomal loci and mediate plasmid and genomic island maintenance through post-segregational killing mechanisms. TA systems exist in surprisingly high numbers in all prokaryotes, but cyanobacterial TA systems have been only very poorly experimentally characterized so far. Cyanobacteria are the only prokaryotes that perform oxygenic photosynthesis. As such, cyanobacteria are of high ecological importance and are considered promising for the production of biofuels. Here, we present the molecular characterization of the sll7003/ssl7004 TA system encoded on plasmid pSYSA of the model cyanobacterium Synechocystis sp. PCC 6803 as involving a Mg2+-dependent RNA endonuclease activity targeting single-stranded RNA regions and demonstrate the functionality of four more TA systems encoded on this 100,749 bp plasmid. Furthermore, one additional type I, one additional type II and three free-standing TA system components are predicted on pSYSA, all of which appear active judged by their expression. By harboring at least seven simultaneously active TA systems, pSYSA appears as the plasmid most strongly selected for among all plasmids studied in this respect thus far. These results point to a high biological relevance of pSYSA, whose coding capacity is to 75% devoted to three distinct CRISPR systems mediating antiviral defense.

Concepts: DNA, Cyanobacteria, Gene, Genetics, Bacteria, Molecular biology, Genome, Synechocystis

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In cyanobacteria, nitrogen homeostasis is maintained by an intricate regulatory network around transcription factor NtcA. Although mechanisms controlling NtcA activity appear to be well understood, its regulon remains poorly defined. To determine the NtcA regulon during the early stages of nitrogen starvation for the model cyanobacterium Synechocystis sp. PCC 6803, we performed chromatin immunoprecipitation, followed by sequencing (ChIP-seq), in parallel with transcriptome analysis (RNA-seq). Through combining these methods, we determined 51 genes activated and 28 repressed directly by NtcA. In addition to genes associated with nitrogen and carbon metabolism, a considerable number of genes without current functional annotation were among direct targets providing a rich reservoir for further studies. The NtcA regulon also included eight non-coding RNAs, of which Ncr1071, Syr6 and NsiR7 were experimentally validated, and their putative targets were computationally predicted. Surprisingly, we found substantial NtcA binding associated with delayed expression changes indicating that NtcA can reside in a poised state controlled by other factors. Indeed, a role of PipX as modulating factor in nitrogen regulation was confirmed for selected NtcA-targets. We suggest that the indicated poised state of NtcA enables a more differentiated response to nitrogen limitation and can be advantageous in native habitats of Synechocystis.

Concepts: DNA, Gene, Gene expression, Bacteria, Metabolism, RNA, Chromatin immunoprecipitation, Synechocystis

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Manganese is an essential element required by cyanobacteria, as it is an essential part of the oxygen-evolving center of photosystem II. In the presence of atmospheric oxygen, manganese is present as manganese oxides, which have low solubility and consequently provide low bioavailability. It is unknown if cyanobacteria are able to utilize these manganese sources, and what mechanisms may be employed to do so. Recent evidence suggests that type IV pili in non-photosynthetic bacteria facilitate electron donation to extracellular electron acceptors, thereby enabling metal acquisition. Our present study investigates whether PilA1 (major pilin protein of type IV pili) enables the cyanobacterium Synechocystis PCC 6808 to access to Mn from manganese oxides. We present physiological and spectroscopic data, which indicate that the presence of PilA1 enhances the ability of cyanobacteria to grow on manganese oxides. These observations suggest a role of PilA1-containing pili in cyanobacterial manganese acquisition.

Concepts: Cyanobacteria, Photosynthesis, Oxygen, Bacteria, Chemical element, Metal, Synechocystis, Pilus

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Safe and effective algaecides are needed to control agriculturally and environmentally significant algae species. Four series (6, 10, 17, and 21) of 29 novel 4-aminopyrimidine derivatives were rationally designed and synthesized. A part of 10, 17, and 21 displayed potent inhibition against Escherichia coli pyruvate dehydrogenase complex E1 (E. coli PDHc-E1) (IC50 = 2.12-18.06 μM) and good inhibition against Synechocystis sp. PCC 6803 (EC50 = 0.7-7.1 μM) and Microcystis sp. FACH 905 (EC50 = 3.7-7.6 μM). The algaecidal activity of these compounds positively correlated with their inhibnitio against E. coli PDHc-E1. In particular, 21l and 10b exhibited potent algaecidal activity against PCC 6803 (EC50 = 0.7 and 0.8 μM, respectively), which were 2-fold increase on the potency if compared to copper sulfate (EC50 = 1.8 μM), also showed the best inhibition against cyanobacteria PDHc-E1 (IC50 = 5.10 and 6.06 μM, respectively). 17h and 21e with the best inhibition against E. coli PDHc-E1 were studied by molecular docking, site-directed mutagenesis and enzymatic assays. These results revealed that the improved inhibition of novel inhibitors compared with the lead compound I was due to forming new hydrogen bond with Leu264 at the active site of E. coli PDHc-E1. The results proved a high potential to obtain effective algaecides by rational design of PDHc-E1 inhibitors.

Concepts: Cyanobacteria, Bacteria, Escherichia coli, Enzyme inhibitor, Enterobacteriaceae, Escherichia, Synechocystis, Pyruvate dehydrogenase complex

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Myristyltrimethylammonium bromide (MTAB) is a cationic surfactant used to improve biomass harvesting and pigment extraction form microalgae, but the mechanisms underlying its effectiveness are poorly defined. We document the mechanisms for enhanced harvesting and pigment extraction for the cyanobacterium Synechocystis sp. PCC 6803 using measurements from flow cytometer, zeta potential, release of soluble components, and microscopy. Harvesting efficiency increased as the MTAB/Biomass dose increased from 0 to 40%. A low MTAB dose (≤ 8%) mainly brought about coagulation and flocculation, which led to aggregation that improved harvesting, but 40% MTAB had the highest harvesting efficiency, 62%. Adding MTAB above a MTAB/Biomass dose of 8% also increased cell-membrane permeability, which allowed the solvent (ethyl acetate) to pass into the cells and resulted in a large increase in extraction efficiency of pigments: An MTAB/Biomass ratio of 60% for 180 min achieved the highest extraction efficiencies of chlorophyll and carotenoids, 95% and 91%, respectively. Combining harvesting and extraction performances with results from flow cytometry, zeta potential, release of soluble components, and microscopy lead to the following mechanistic understandings. MTAB dose from 8% to 40% solubilized EPS, which lowered the biomass’s negative charge, but caused breakup of the large aggregates. An increase of cell permeability also in this stage allowed ethyl acetate to pass into the cells and achieve better pigment extraction. MTAB >40% led to cell lysis and a large increase in soluble organics, but complete cell lysis was not required to achieve the maximum extraction efficiency. The MTAB/Biomass % ratio for optimizing harvest efficiency and pigment extraction lay in the range of 40%-60%.

Concepts: Photosynthesis, Cell biology, Flow cytometry, Acetic acid, Surfactant, Fluorescein, Synechocystis, Propidium iodide

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One strategy for enhancing photosynthesis in crop plants is to improve their ability to repair photosystem II (PSII) in response to irreversible damage by light. Despite the pivotal role of thylakoid-embedded FtsH protease complexes in the selective degradation of PSII subunits during repair, little is known about the factors involved in regulating FtsH expression. Here we show using the cyanobacterium Synechocystis sp. PCC 6803 that the Psb29 subunit, originally identified as a minor component of His-tagged PSII preparations, physically interacts with FtsH complexes in vivo and is required for normal accumulation of the FtsH2/FtsH3 hetero-oligomeric complex involved in PSII repair. We show using X-ray crystallography that Psb29 from Thermosynechococcus elongatus has a unique fold consisting of a helical bundle and an extended C-terminal helix and contains a highly conserved region that might be involved in binding to FtsH. A similar interaction is likely to occur in Arabidopsis chloroplasts between the Psb29 homologue, termed THF1, and the FTSH2/FTSH5 complex. The direct involvement of Psb29/THF1 in FtsH accumulation helps explain why THF1 is a target during the hypersensitive response in plants induced by pathogen infection. Downregulating FtsH function and the PSII repair cycle via THF1 would contribute to the production of reactive oxygen species, the loss of chloroplast function and cell death.This article is part of the themed issue ‘Enhancing photosynthesis in crop plants: targets for improvement’.

Concepts: Cyanobacteria, Photosynthesis, Oxygen, Eukaryote, Chloroplast, Plant, Photosystem, Synechocystis