Cyanobacterium Anabaena Sp Research Articles

Structure Insights Into Photosystem I Octamer From Cyanobacteria

The diversity of photosystem oligomers is essential to understanding how photosynthetic organisms adapt to light conditions. Due to its structural and physiological significance, the assembly of the PSI supercomplex has been of great interest recently in terms of both chloroplast and cyanobacteria. In this study, two novel photosystem I supercomplexes were isolated for the first time from the low light incubated culture of filamentous cyanobacterium Anabaena sp. PCC 7120. These complexes were defined as PSI hexamers and octamers through biochemical and biophysical characterization. Their 77K emission spectra indicated that the red forms of chlorophylls seemed not to be affected during oligomerization. By cryo-EM single-particle analysis, a near-atomic (7.0 Å) resolution structure of a PSI octamer was resolved, and the molecular assemblies of a stable PSI octamer were revealed.

The Importance of the C-Terminal Cys Pair of Phosphoribulokinase in Phototrophs in Thioredoxin-Dependent Regulation.

Phosphoribulokinase (PRK), one of the enzymes in the Calvin–Benson cycle, is a well-known target of thioredoxin (Trx), which regulates various enzyme activities by the reduction of disulfide bonds in a light-dependent manner. PRK has two Cys pairs conserved in the N-terminal and C-terminal regions, and the N-terminal one near the active site is thought to be responsible for the regulation. The flexible clamp loop located between the N-terminal two Cys residues has been deemed significant to Trx-mediated regulation. However, cyanobacterial PRK is also subject to Trx-dependent activation despite the lack of this clamp loop. We, therefore, compared Trx-mediated regulation of PRK from the cyanobacterium Anabaena sp. PCC 7120 (A.7120_PRK) and that from the land plant Arabidopsis thaliana (AtPRK). Interestingly, peptide mapping and site-directed mutagenesis analysis showed that Trx was more effective in changing the redox states of the C-terminal Cys pair in both A.7120_PRK and AtPRK. In addition, the effect of redox state change of the C-terminal Cys pair on PRK activity was different between A.7120_PRK and AtPRK. Trx-mediated redox regulation of the C-terminal Cys pair was also important for complex dissociation/formation with CP12 and glyceraldehyde 3-phosphate dehydrogenase. Furthermore, in vivo analysis of the redox states of PRK showed that only one disulfide bond is reduced in response to light. Based on the enzyme activity assay and the complex formation analysis, we concluded that Trx-mediated regulation of the C-terminal Cys pair of PRK is important for activity regulation in cyanobacteria and complex dissociation/formation in both organisms.

A tRNA t6A modification system contributes to the sensitivity towards the toxin β-N-methylamino-L-alanine (BMAA) in the cyanobacterium Anabaena sp. PCC 7120.

Cyanobacteria are oxygen-evolving photosynthetic autotrophs essential for nutrient cycling in the environment. They possess multiple control mechanisms for their cellular activities in order to adapt to the environment. While protein translation is essential for cell survival and adaptation, the regulation and the flexibility of this process are poorly understood in cyanobacteria. β-N-methylamino-L-alanine (BMAA), an amino acid analog proposed as an environmental neurotoxin, is highly toxic to the filamentous diazotrophic cyanobacterium Anabaena PCC 7120. In this study, through genetic analysis of BMAA-resistant mutants, we demonstrate that the system responsible for modification of ANN-decoding tRNAs with N(6)-threonylcarbamoyl adenosine (t6A) is involved in BMAA sensitivity through the control of translation. Both BMAA and inactivation of the t6A biosynthesis pathway affect translational fidelity and ribosome assembly. However, the two factors display either additive effects on translational elongation, or attenuate each other over translational fidelity or the resistance/sensitivity to antibiotics that inhibit different steps of the translational process. BMAA has a broad effect on translation and transcription, and once BMAA enters the cells, the presence of the t6A biosynthesis pathway increases the sensitivity of the cells towards this toxin. BMAA-resistant mutants screening is an effective method for getting insight into the toxic mechanisms of BMAA. In addition, BMAA is a useful tool for probing translational flexibility of cyanobacteria, and the characterization of the corresponding resistant mutants should help us to reveal translational mechanism allowing cyanobacteria to adapt to changing environments.

Cyclophilin anaCyp40 regulates photosystem assembly and phycobilisome association in a cyanobacterium

Cyclophilins, or immunophilins, are proteins found in many organisms including bacteria, plants and humans. Most of them display peptidyl-prolyl cis-trans isomerase activity, and play roles as chaperones or in signal transduction. Here, we show that cyclophilin anaCyp40 from the cyanobacterium Anabaena sp. PCC 7120 is enzymatically active, and seems to be involved in general stress responses and in assembly of photosynthetic complexes. The protein is associated with the thylakoid membrane and interacts with phycobilisome and photosystem components. Knockdown of anacyp40 leads to growth defects under high-salt and high-light conditions, and reduced energy transfer from phycobilisomes to photosystems. Elucidation of the anaCyp40 crystal structure at 1.2-Å resolution reveals an N-terminal helical domain with similarity to PsbQ components of plant photosystem II, and a C-terminal cyclophilin domain with a substrate-binding site. The anaCyp40 structure is distinct from that of other multi-domain cyclophilins (such as Arabidopsis thaliana Cyp38), and presents features that are absent in single-domain cyclophilins.

Regulatory role of LexA in modulating photosynthetic redox poise and cadmium stress tolerance in the cyanobacterium, Anabaena sp. PCC7120

Strategies developed by organisms to overcome disruption in redox poise of photosynthetic electron transport chain (pETC) are important for its survival under abiotic stress. The process needs to be tightly regulated for optimal functioning. While the redox poising processes are well known in cyanobacteria, understanding of their regulatory network is lacking. Since LexA is one of the known global regulators of stress response in the cyanobacterium Anabaena sp. PCC7120, its role in pETC redox poising was investigated using cadmium (Cd) as an abiotic stressor to disrupt photosynthesis. Assessment of the photosynthetic responses of recombinant Anabaena strains, An lexA + (LexA-overexpressing) and AnpAM (vector control), under unstressed and Cd-stressed conditions using transmission electron microscopy (TEM) and chlorophyll a fluorescence, indicated that some pETC redox poising responses, including PSII photodamage, energy dissipation, PSI photoprotection, and NDH-mediated cyclic electron flow were decreased in An lexA + under unstressed conditions. Disturbance in pETC redox poise during Cd stress observed in Anabaena was accentuated upon overexpression of LexA. The decreased photodamage of PSII and increased photoinhibition of PSI in An lexA + in the presence or absence of Cd stress, correlated well with the changes in pETC complexes observed in blue native (BN)-PAGE and the regulation of over 70 of the 90 pETC component genes by LexA demonstrated through transcript, electromobility shift assay (EMSA), and bioinformatics studies. In a nutshell, LexA has been identified as one of the regulators involved in the streamlining of pETC redox poising responses under normal growth and during abiotic stress through transcriptional regulation of some of the redox-controlled pETC component genes. • Abiotic stresses disrupt photosynthetic redox poise, triggering poising responses. • Poising responses are known in cyanobacteria, but their regulatory network is not. • LexA overexpression in Anabaena reduced these responses, lowering stress tolerance. • In-silico , qRT-PCR and EMSA showed LexA regulation on over 70 photosynthetic genes. • LexA was shown to modulate these responses by regulating some photosynthetic genes.

Grazer-induced changes in molecular signatures of cyanobacteria

Algal biomass production is an emerging renewable source of fuels, nutrients, manufacturing materials, and pharmaceuticals. Industrial-scale production is predominantly performed in open raceway ponds that are inexpensive to build, operate, and maintain compared to closed bioreactors. However, these open pond systems suffer from increased opportunities for biological contamination from predators, pathogens, and competitors, which result in reduced biomass quality and yields and often the complete destruction of the crop over a short period of time. Early detection of contaminants is a necessary step of integrated pest management for triggering and informing interventions to prevent crop losses. To develop a sensitive method of detection utilizing mass spectrometry (MS), we used three methods – imaging mass spectrometry (imaging MS), liquid chromatography MS/MS (LC-MS/MS) combined with molecular networking, and gas chromatography MS/MS (GC–MS/MS) – to observe and identify molecular signatures from a model predator-prey system of the heterolobosean amoeba HGG1 preying on the filamentous cyanobacterium Anabaena sp. PCC 7120. Imaging MS enabled the association of molecules with the crop, the predator, or the activity of protozoan grazing, while LC-MS/MS-based molecular networking identified a subset of the grazing-specific signals as the chlorophyll breakdown products pyropheophytin, pheophorbide A, and pyropheophorbide. Application of MS techniques to other amoeba-cyanobacterial predator-prey pairs allowed categorization of grazing signatures as universal or specific to the species of prey, predator, or the prey-predator pairing, indicating that MS-based techniques can distinguish crops from competitors and potentially identify predators. Finally, GCMS/MS was shown to be capable of monitoring novel volatile organic compounds (VOCs), including those predicted to be released through chlorophyll breakdown, in the headspace over algal cultures under predation. These results demonstrate that the combination of multiple MS technologies creates a predictive framework to identify and catalog relevant molecular signatures for informing crop protection strategies.

Excitation-energy transfer in heterocysts isolated from the cyanobacterium Anabaena sp. PCC 7120 as studied by time-resolved fluorescence spectroscopy

Heterocysts are formed in filamentous heterocystous cyanobacteria under nitrogen-starvation conditions, and possess a very low amount of photosystem II (PSII) complexes than vegetative cells. Molecular, morphological, and biochemical characterizations of heterocysts have been investigated; however, excitation-energy dynamics in heterocysts are still unknown. In this study, we examined excitation-energy-relaxation processes of pigment-protein complexes in heterocysts isolated from the cyanobacterium Anabaena sp. PCC 7120. Thylakoid membranes from the heterocysts showed no oxygen-evolving activity under our experimental conditions and no thermoluminescence-glow curve originating from charge recombination of S2QA−. Two dimensional blue-native/SDS-PAGE analysis exhibits tetrameric, dimeric, and monomeric photosystem I (PSI) complexes but almost no dimeric and monomeric PSII complexes in the heterocyst thylakoids. The steady-state fluorescence spectrum of the heterocyst thylakoids at 77 K displays both characteristic PSI fluorescence and unusual PSII fluorescence different from the fluorescence of PSII dimer and monomer complexes. Time-resolved fluorescence spectra at 77 K, followed by fluorescence decay-associated spectra, showed different PSII and PSI fluorescence bands between heterocysts and vegetative thylakoids. Based on these findings, we discuss excitation-energy-transfer mechanisms in the heterocysts.

Proteomics analysis reveals several metabolic alterations in cyanobacterium Anabaena sp. NC-K1 in response to alpha-cypermethrin exposure.

In the current study, the effect of the EC50 and LC90 concentrations of pyrethroid insecticide alpha-cypermethrin to cyanobacteria Anabaena sp. NC-K1 was investigated at different time exposures (1st day, 4th day and 7th day) with reference to growth, photosynthetic pigments, oxidative damage and antioxidant defence system. Superoxide dismutase (1.38-fold), peroxidase (5.04) and proline content (2.27-fold) were enhanced compared to the control. After performing 2D gel electrophoresis at 1st day EC50 exposure, where appropriate differences in the biochemical and physiological parameters were observed, 22 differentially accumulated proteins (20 upregulated and 2 downregulated) were selected for mass spectrometry. Out of 42 proteins identified, 20 upregulated protein spots were classified into twelve categories according to their metabolic functions. Proteins related to photosynthesis (phycobilisome rod-core linker polypeptide, rubisco), stress responses (Hsp70, Hsp40, catalase family peroxidase), translation (elongation factor Tu) and amino acid biosynthesis and metabolism (3-phosphoshikimate 1-carboxyvinyl transferase) were significantly upregulated. Additionally, proteins involved in transcription and DNA repair (Snf-2 histone linker phd ring helicase, RNA polymerase sigma factor RpoD and Holliday junction ATP-dependent DNA helicase RuvA) were considerably upregulated. Upregulation of these proteins against pesticide stress presumably maintained the photosynthesis, energy metabolism, carbohydrate metabolism, transport and signalling proteins, transcription, translation and DNA repair. Additionally, these proteins might involve in sufficient detoxification of ROS and play a crucial role in damage removal and repair of oxidized proteins, lipids and nucleic acids. Taken together, Anabaena sp. NC-K1 responded towards alpha-cypermethrin stress via modulating its proteome to maintain its cellular metabolism and homeostasis.

Functions of the Essential Gene mraY in Cellular Morphogenesis and Development of the Filamentous Cyanobacterium Anabaena PCC 7120.

Bacterial cell shape is determined by the peptidoglycan (PG) layer. The cyanobacterium Anabaena sp. PCC 7120 (Anabaena) is a filamentous strain with ovoid-shaped cells connected together with incomplete cell constriction. When deprived of combined nitrogen in the growth medium, about 5–10% of the cells differentiate into heterocysts, cells devoted to nitrogen fixation. It has been shown that PG synthesis is modulated during heterocyst development and some penicillin-binding proteins (PBPs) participating in PG synthesis are required for heterocyst morphogenesis or functioning. Anabaena has multiple PBPs with functional redundancy. In this study, in order to examine the function of PG synthesis and its relationship with heterocyst development, we created a conditional mutant of mraY, a gene necessary for the synthesis of the PG precursor, lipid I. We show that mraY is required for cell and filament integrity. Furthermore, when mraY expression was being limited, persistent septal PG synthetic activity was observed, resulting in increase in cell width. Under non-permissive conditions, filaments and cells were rapidly lysed, and no sign of heterocyst development within the time window allowed was detected after nitrogen starvation. When mraY expression was being limited, a high percentage of heterocyst doublets were found. These doublets are formed likely as a consequence of delayed cell division and persistent septal PG synthesis. MraY interacts with components of both the elongasome and the divisome, in particular those directly involved in PG synthesis, including HetF, which is required for both cell division and heterocyst formation.

Single-Cell Measurements of Fixation and Intercellular Exchange of C and N in the Filaments of the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120.

ABSTRACTUnder diazotrophic conditions, the model filamentous, heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120 develops a metabolic strategy based on the physical separation of the processes of oxygenic photosynthesis, in vegetative cells, and N2 fixation, in heterocysts. This strategy requires the exchange of carbon and nitrogen metabolites and their distribution along the filaments, which takes place through molecular diffusion via septal junctions involving FraCD proteins. Here, Anabaena was incubated in a time course (up to 20 h) with [13C]bicarbonate and 15N2 and analyzed by secondary ion mass spectrometry imaging (SIMS) (large-geometry SIMS [LG-SIMS] and NanoSIMS) to quantify C and N assimilation and distribution in the filaments. The 13C/12C and 15N/14N ratios measured in wild-type filaments showed a general increase with time. The enrichment was relatively homogeneous in vegetative cells along individual filaments, while it was reduced in heterocysts. Heterocysts, however, accumulated recently fixed N at their poles, in which the cyanophycin plug [multi-l-arginyl-poly(l-aspartic acid)] is located. In contrast to the rather homogeneous label found along stretches of vegetative cells, 13C/12C and 15N/14N ratios were significantly different between filaments both at the same and different time points, showing high variability in metabolic states. A fraC fraD mutant did not fix N2, and the 13C/12C ratio was homogeneous along the filament, including the heterocyst in contrast to the wild type. Our results show the consumption of reduced C in the heterocysts associated with the fixation and export of fixed N and present an unpredicted heterogeneity of cellular metabolic activity in different filaments of an Anabaena culture under controlled conditions.

Role of a cryptic tRNA gene operon in survival under translational stress.

As compared to eukaryotes, bacteria have a reduced tRNA gene set encoding between 30 and 220 tRNAs. Although in most bacterial phyla tRNA genes are dispersed in the genome, many species from distinct phyla also show genes forming arrays. Here, we show that two types of arrays with distinct evolutionary origins exist. This work focuses on long tRNA gene arrays (L-arrays) that encompass up to 43 genes, which disseminate by horizontal gene transfer and contribute supernumerary tRNA genes to the host. Although in the few cases previously studied these arrays were reported to be poorly transcribed, here we show that the L-array of the model cyanobacterium Anabaena sp. PCC 7120, encoding 23 functional tRNAs, is largely induced upon impairment of the translation machinery. The cellular response to this challenge involves a global reprogramming of the transcriptome in two phases. tRNAs encoded in the array are induced in the second phase of the response, directly contributing to cell survival. Results presented here show that in some bacteria the tRNA gene set may be partitioned between a housekeeping subset, which constantly sustains translation, and an inducible subset that is generally silent but can provide functionality under particular conditions.

Comparative Phenotypic Analysis of Anabaena sp. PCC 7120 Mutants of Porinlike Genes.

Porins are essential for the viability of Gram-negative bacteria. They ensure the uptake of nutrients, can be involved in the maintenance of outer membrane integrity and define the antibiotic or drug resistance of organisms. The function and structure of porins in proteobacteria is well described, while their function in photoautotrophic cyanobacteria has not been systematically explored. We compared the domain architecture of nine putative porins in the filamentous cyanobacterium Anabaena sp. PCC 7120 and analyzed the seven candidates with predicted OprB-domain. Single recombinant mutants of the seven genes were created and their growth capacity under different conditions was analyzed. Most of the putative porins seem to be involved in the transport of salt and copper, as respective mutants were resistant to elevated concentrations of these substances. In turn, only the mutant of alr2231 was less sensitive to elevated zinc concentrations, while mutants of alr0834, alr4741 and all4499 were resistant to high manganese concentrations. Notably the mutant of alr4550 shows a high sensitivity against harmful compounds, which is indicative for a function related to the maintenance of outer membrane integrity. Moreover, the mutant of all5191 exhibited a phenotype which suggests either a higher nitrate demand or an inefficient nitrogen fixation. The dependency of porin membrane insertion on Omp85 proteins was tested exemplarily for Alr4550, and an enhanced aggregation of Alr4550 was observed in two omp85 mutants. The comparative analysis of porin mutants suggests that the proteins in parts perform distinct functions related to envelope integrity and solute uptake.

Physiological and Proteomic Studies of the Cyanobacterium Anabaena sp. Acclimated to Desiccation Stress.

Agricultural productivity is threatened by increasing incidence of drought and the drought tolerant cyanobacteria offer a better solution in the restoration of soil fertility and productivity. The present study describes the comparative physiological response of the cyanobacterium Anabaena sp. acclimated and un-acclimated to desiccation stress induced by polyethylene glycol (10% PEG). While, the acclimated cyanobacterial cells grew luxuriantly with optimal chlorophyll content, photosynthetic activities and nitrogen fixation, the un-acclimated cells exhibited reduced growth rate, chlorophyll content, photosynthetic activities and nitrogen fixation. Distinct differences in the accumulation of lipid peroxidation products, proline and activity of superoxide dismutase were observed under identical growth conditions in the acclimated and un-acclimated cells. Desiccation-acclimated and un-acclimated cyanobacteria showed significant alterations in the abundance of important proteins in the proteome. Two-dimensional gel electrophoresis followed by MALDI-TOF-MS/MS analysis identified twelve proteins. The acclimated cells showed the up regulation of proteins such as Rubisco, fructose-bis-phosphate aldolase, fructose 1-6 bisphosphatase, phosphoglycerate dehydrogenase and elongation factors Tu and Ts as compared to un-acclimated cells. Therefore, the ability to maintain photosynthesis, antioxidants and increased accumulation of proteins related to energy metabolism helped the acclimated cyanobacterium Anabaena sp. to grow optimally under desiccation stress conditions.

Calm on the surface, dynamic on the inside. Molecular homeostasis of Anabaena sp. PCC 7120 nitrogen metabolism.

Nitrogen sources are all converted into ammonium/ia as a first step of assimilation. It is reasonable to expect that molecular components involved in the transport of ammonium/ia across biological membranes connect with the regulation of both nitrogen and central metabolism. We applied both genetic (i.e., Δamt mutation) and environmental treatments to a target biological system, the cyanobacterium Anabaena sp PCC 7120. The aim was to both perturb nitrogen metabolism and induce multiple inner nitrogen states, respectively, followed by targeted quantification of key proteins, metabolites and enzyme activities. The absence of AMT transporters triggered a substantial whole-system response, affecting enzyme activities and quantity of proteins and metabolites, spanning nitrogen and carbon metabolisms. Moreover, the Δamt strain displayed a molecular fingerprint indicating nitrogen deficiency even under nitrogen replete conditions. Contrasting with such dynamic adaptations was the striking near-complete lack of an externally measurable altered phenotype. We conclude that this species evolved a highly robust and adaptable molecular network to maintain homeostasis, resulting in substantial internal but minimal external perturbations. This analysis provides evidence for a potential role of AMT transporters in the regulatory/signalling network of nitrogen metabolism and the existence of a novel fourth regulatory mechanism controlling glutamine synthetase activity.

Disentangling the Impact of Sulfur Limitation on Exopolysaccharide and Functionality of Alr2882 by In Silico Approaches in Anabaena sp. PCC 7120

The wide applications, uniqueness, and high quality of cyanobacterial exopolysaccharides (EPSs) have attracted many biotechnologists. Despite it, the inducers and molecular determinants of EPS biosynthesis in cyanobacteria are lesser known. Although, studies revealed that environmental cues especially C/N ratio as the prime modulator, the factors like light, temperature, moisture, and nutrient availability, etc. have been overlooked. Due to this, the possibilities to modify cyanobacterial system for achieving higher quantity of EPS either by modifying growth medium or metabolic engineering are restricted to few optimisations. Therefore, the present work describes the impact of sulfate limitations on the EPS production and compositions in the cyanobacterium Anabaena sp. PCC 7120. Increased EPS production with enhanced expression of alr2882 was observed in lower sulfate supplementations; however, FTIR analysis depicted an altered composition of supramolecule. Furthermore, in silico analysis of Alr2882 depicted the presence of ExoD domain and three transmembrane regions, thereby indicating its membrane localisation and role in the EPS production. Additionally, the phylogeny and multiple sequence alignment showed vertical inheritance of exoD and conservation among cyanobacteria. The meta-threading template-based modelling and ab initio full atomic relaxation by LOMET and ModRefiner servers, respectively, also exhibited helical topology of Alr2882, with nine α-helices arranged antiparallel to the preceding one. Moreover, post-translational modifications predicted in Alr2882 indicated high order of molecular regulation underlining EPS production in Anabaena sp. PCC 7120. This study provides a foundation for understanding the EPS biosynthesis mechanism under sulfur limitation and the possible role of ExoD in cyanobacteria.

Two novel heteropolymer-forming proteins maintain the multicellular shape of the cyanobacterium Anabaena sp. PCC 7120.

Polymerizing and filament-forming proteins are instrumental for numerous cellular processes such as cell division and growth. Their function in stabilization and localization of protein complexes and replicons is achieved by a filamentous structure. Known filamentous proteins assemble into homopolymers consisting of single subunits - for example, MreB and FtsZ in bacteria - or heteropolymers that are composed of two subunits, for example, keratin and α/β tubulin in eukaryotes. Here, we describe two novel coiled-coil-rich proteins (CCRPs) in the filament-forming cyanobacterium Anabaena sp. PCC 7120 (hereafter Anabaena) that assemble into a heteropolymer and function in the maintenance of the Anabaena multicellular shape (termed trichome). The two CCRPs - Alr4504 and Alr4505 (named ZicK and ZacK) - are strictly interdependent for the assembly of protein filaments invivo and polymerize nucleotide independently invitro, similar to known intermediate filament (IF) proteins. A ΔzicKΔzacK double mutant is characterized by a zigzagged cell arrangement and hence a loss of the typical linear Anabaena trichome shape. ZicK and ZacK interact with themselves, with each other, with the elongasome protein MreB, the septal junction protein SepJ and the divisome associate septal protein SepI. Our results suggest that ZicK and ZacK function in cooperation with SepJ and MreB to stabilize the Anabaena trichome and are likely essential for the manifestation of the multicellular shape in Anabaena. Our study reveals the presence of filament-forming IF-like proteins whose function is achieved through the formation of heteropolymers in cyanobacteria.

Inactivation of Three RG(S/T)GR Pentapeptide-Containing Negative Regulators of HetR Results in Lethal Differentiation of Anabaena PCC 7120.

The filamentous cyanobacterium Anabaena sp. PCC 7120 produces, during the differentiation of heterocysts, a short peptide PatS and a protein HetN, both containing an RGSGR pentapeptide essential for activity. Both act on the master regulator HetR to guide heterocyst pattern formation by controlling the binding of HetR to DNA and its turnover. A third small protein, PatX, with an RG(S/T)GR motif is present in all HetR-containing cyanobacteria. In a nitrogen-depleted medium, inactivation of patX does not produce a discernible change in phenotype, but its overexpression blocks heterocyst formation. Mutational analysis revealed that PatX is not required for normal intercellular signaling, but it nonetheless is required when PatS is absent to prevent rapid ectopic differentiation. Deprivation of all three negative regulators—PatS, PatX, and HetN—resulted in synchronous differentiation. However, in a nitrogen-containing medium, such deprivation leads to extensive fragmentation, cell lysis, and aberrant differentiation, while either PatX or PatS as the sole HetR regulator can establish and maintain a semiregular heterocyst pattern. These results suggest that tight control over HetR by PatS and PatX is needed to sustain vegetative growth and regulated development. The mutational analysis has been interpreted in light of the opposing roles of negative regulators of HetR and the positive regulator HetL.

Response of a rice-field cyanobacterium Anabaena sp. HKAR-7 upon exposure to ultraviolet-B radiation and ammonium chloride

Interactive effects of ultraviolet radiation (UVR), photosynthetically active radiation (PAR) and exogenously supplied ammonium chloride (NH4Cl) was studied in the rice-field cyanobacterium Anabaena sp. HKAR-7. The cyanobacterium was cultured under varying NH4Cl concentrations i.e., 0, 50, 200, 500, 1000 and 5000 μM and 200 μM (concentration) was found to be optimum for the growth of the cyanobacterium. Detrimental effects of UV-B exposure were observed on photosynthetic pigments such as chlorophyll a (Chl a), carotenoids and phycocyanin (PC). However, damage to these pigments was less in the cyanobacterial samples supplemented with NH4Cl. Contents of Chl a and PC in cyanobacterial cells decreased upon UV-B exposure but decrement was less in the samples supplemented with NH4Cl. Upon UV-B exposure, carotenoids content enhanced initially (till 15 days) during the course of treatment (21 days) but significant decrease (in carotenoids content) was observed in later phase of the experiment. From the results of photosynthetic activity, maximum quantum efficiency of PSII (Fv/Fm) and maximum electron transport rate (ETRmax), it could be concluded that exogenous supplementation of NH4Cl (optimum concentration) helped in protecting the cyanobacterial cells from highly energetic UVR to certain extent. Another interesting observation was significantly higher levels of biosynthesis and accumulation of mycosporine-like amino acids (MAAs) in the cyanobacterial cells supplemented with NH4Cl in comparison to non-supplemented cells. The purified MAA was identified to be phorphyra-334 as evidenced by UV/VIS absorption spectra, high performance liquid chromatography (HPLC) and electrospray ionization-mass spectrometry (ESI–MS).

Microplastics as vectors of the antibiotics azithromycin and clarithromycin: Effects towards freshwater microalgae

Water pollution due to microplastics (MPs) is recognized as a major anthropogenic impact. Once MPs reach the ecosystems, they are exposed to a variety of other pollutants, which can be sorbed on them, transported and eventually desorbed. In this work, we tested the hypothesis that MPs can behave as conveyors for delivering chemicals toxic to aquatic microorganisms by investigating the vector role of MPs of polyethylene terephthalate (PET), polylactic acid (PLA), polyoxymethylene (POM) and polystyrene (PS) to the macrolide antibiotics azithromycin (AZI) and clarithromycin (CLA). AZI and CLA were chosen, as they are included in the Watch List for EU monitoring concerning water policy by Decision (EU) 2018/840. MPs were loaded in contact with 500 μg/L of AZI or 1000 μg/L of CLA. Results showed that both antibiotics were sorbed on all tested MPs. The more hydrophobic AZI was sorbed in higher proportion than CLA. Both antibiotics were desorbed from MPs upon contact with water with percentages between 14.6 ± 2.6% for AZI and 1.9 ± 1.4% for CLA of the concentrations to which the MPs were initially exposed. Virgin MPs were not toxic to the cyanobacterium Anabaena sp. PCC7120. However, antibiotic-loaded MPs significantly inhibited the growth and chlorophyll content of the cyanobacterium. Most of the sorbed antibiotics became released upon contact with cyanobacterial cultures, which was the cause for the observed toxicity. Therefore, MPs can play a role as vectors of antibiotics in freshwaters systems affecting the basic trophic level of photosynthetic microorganisms.

The Inorganic Nutrient Regime and the mre Genes Regulate Cell and Filament Size and Morphology in the Phototrophic Multicellular Bacterium Anabaena.

The model cyanobacterium Anabaena sp. PCC 7120 exhibits a phototrophic metabolism relying on oxygenic photosynthesis and a complex morphology. The organismic unit is a filament of communicated cells that may include cells specialized in different nutritional tasks, thus representing a paradigm of multicellular bacteria. In Anabaena, the inorganic carbon and nitrogen regime influenced not only growth, but also cell size, cell shape, and filament length, which also varied through the growth cycle. When using combined nitrogen, especially with abundant carbon, cells enlarged and elongated during active growth. When fixing N2, which imposed lower growth rates, shorter and smaller cells were maintained. In Anabaena, gene homologs to mreB, mreC, and mreD form an operon that was expressed at higher levels during the phase of fastest growth. In an ntcA mutant, mre transcript levels were higher than in the wild type and, consistently, cells were longer. Negative regulation by NtcA can explain that Anabaena cells were longer in the presence of combined nitrogen than in diazotrophic cultures, in which the levels of NtcA are higher. mreB, mreC, and mreD mutants could grow with combined nitrogen, but only the latter mutant could grow diazotrophically. Cells were always larger and shorter than wild-type cells, and their orientation in the filament was inverted. Consistent with increased peptidoglycan width and incorporation in the intercellular septa, filaments were longer in the mutants, suggesting a role for MreB, MreC, and MreD in the construction of septal peptidoglycan that could affect intercellular communication required for diazotrophic growth.IMPORTANCE Most studies on the determination of bacterial cell morphology have been conducted in heterotrophic organisms. Here, we present a study of how the availability of inorganic nitrogen and carbon sources influence cell size and morphology in the context of a phototrophic metabolism, as found in the multicellular cyanobacterium Anabaena In Anabaena, the expression of the MreB, MreC, and MreD proteins, which influence cell size and length, are regulated by NtcA, a transcription factor that globally coordinates cellular responses to the C-to-N balance of the cells. Moreover, MreB, MreC, and MreD also influence septal peptidoglycan construction, thus affecting filament length and, possibly, intercellular molecular exchange that is required for diazotrophic growth. Thus, here we identified new roles for Mre proteins in relation to the phototrophic and multicellular character of a cyanobacterium, Anabaena.