Search Results (7)

Non-proteinogenic neurotoxic amino acid β-N-methylamino-L-alanine (BMAA) is synthesized by cyanobacteria, diatoms, and dinoflagellates, and is known to be a causative agent of human neurodegenerative diseases. Different phytoplankton organisms’ ability to synthesize BMAA could indicate the importance of this molecule in the interactions between microalgae in nature. We were interested in the following: what kinds of mechanisms underline BMAA’s action on cyanobacterial cells in different nitrogen supply conditions. Herein, we present a proteomic analysis of filamentous cyanobacteria Nostoc sp. PCC 7120 cells that underwent BMAA treatment in diazotrophic conditions. In diazotrophic growth conditions, to survive, cyanobacteria can use only biological nitrogen fixation to obtain nitrogen for life. Note that nitrogen fixation is an energy-consuming process. In total, 1567 different proteins of Nostoc sp. PCC 7120 were identified by using LC-MS/MS spectrometry. Among them, 123 proteins belonging to different functional categories were selected—due to their notable expression differences—for further functional analysis and discussion. The presented proteomic data evidences that BMAA treatment leads to very strong (up to 80%) downregulation of α (NifD) and β (NifK) subunits of molybdenum-iron protein, which is known to be a part of nitrogenase. This enzyme is responsible for catalyzing nitrogen fixation. The genes nifD and nifK are under transcriptional control of a global nitrogen regulator NtcA. In this study, we have found that BMAA impacts in a total of 22 proteins that are under the control of NtcA. Moreover, BMAA downregulates 18 proteins that belong to photosystems I or II and light-harvesting complexes; BMAA treatment under diazotrophic conditions also downregulates five subunits of ATP synthase and enzyme NAD(P)H-quinone oxidoreductase. Therefore, we can conclude that the disbalance in energy and metabolite amounts leads to severe intracellular stress that induces the upregulation of stress-activated proteins, such as starvation-inducible DNA-binding protein, four SOS-response enzymes, and DNA repair enzymes, nine stress-response enzymes, and four proteases. The presented data provide new leads into the ecological impact of BMAA on microalgal communities that can be used in future investigations. Full article

The pentapeptide repeat protein (PRP) superfamily, identified in 1998, has grown to nearly 39,000 sequences from over 3300 species. PRPs, recognized as having at least eight contiguous pentapeptide repeats (PRs) of a consensus pentapeptide sequence, adopt a remarkable structure, namely, a right-handed quadrilateral β-helix with four consecutive PRs forming a single β-helix coil. Adjacent coils join together to form a β-helix “tower” stabilized by β-ladders on the tower faces and type I, type II, or type IV β-turns facilitating an approximately −90° redirection of the polypeptide chain joining one coil face to the next. PRPs have been found in all branches of life, but they are predominantly found in cyanobacteria. Cyanobacteria have existed on earth for more than two billion years and are thought to be responsible for oxygenation of the earth’s atmosphere. Filamentous cyanobacteria such as Nostoc sp. strain PCC 7120 may also represent the oldest and simplest multicellular organisms known to undergo cell differentiation on earth. Knowledge of the biochemical function of these PRPs is essential to understanding how ancient cyanobacteria achieved functions critical to early development of life on earth. PRPs are predicted to exist in all cyanobacteria compartments including thylakoid and cell-wall membranes, cytoplasm, and thylakoid periplasmic space. Despite their intriguing structure and importance to understanding ancient cyanobacteria, the biochemical functions of PRPs in cyanobacteria remain almost completely unknown. The precise biochemical function of only a handful of PRPs is currently known from any organisms, and three-dimensional structures of only sixteen PRPs or PRP-containing multidomain proteins from any organism have been reported. In this review, the current knowledge of the structures and functions of PRPs is presented and discussed. Full article

Nostoc (Anabaena) sp. PCC 7120 is a filamentous cyanobacterial species that fixes N₂ to nitrogenous compounds using specialised heterocyst cells. Changes in the intracellular ratio of carbon to nitrogen (C/N balance) is known to trigger major transcriptional reprogramming of the cell, including initiating the differentiation of vegetative cells to heterocysts. Substantial transcriptional analysis has been performed on Nostoc sp. PCC 7120 during N stepdown (low to high C/N), but not during C stepdown (high to low C/N). In the current study, we shifted the metabolic balance of Nostoc sp. PCC 7120 cultures grown at 3% CO₂ by introducing them to atmospheric conditions containing 0.04% CO₂ for 1 h, after which the changes in gene expression were measured using RNAseq transcriptomics. This analysis revealed strong upregulation of carbon uptake, while nitrogen uptake and metabolism and early stages of heterocyst development were downregulated in response to the shift to low CO₂. Furthermore, gene expression changes revealed a decrease in photosynthetic electron transport and increased photoprotection and reactive oxygen metabolism, as well a decrease in iron uptake and metabolism. Differential gene expression was largely attributed to change in the abundances of the metabolites 2-phosphoglycolate and 2-oxoglutarate, which signal a rapid shift from fluent photoassimilation to glycolytic metabolism of carbon after transition to low CO₂. This work shows that the C/N balance in Nostoc sp. PCC 7120 rapidly adjusts the metabolic strategy through transcriptional reprogramming, enabling survival in the fluctuating environment. Full article

All cyanobacteria produce a neurotoxic non-protein amino acid β-N-methylamino-L-alanine (BMAA). However, the biological function of BMAA in the regulation of cyanobacteria metabolism still remains undetermined. It is known that BMAA suppresses the formation of heterocysts in diazotrophic cyanobacteria under nitrogen starvation conditions, and BMAA induces the formation of heterocyst-like cells under nitrogen excess conditions, by causing the expression of heterocyst-specific genes that are usually “silent” under nitrogen-replete conditions, as if these bacteria receive a nitrogen deficiency intracellular molecular signal. In order to find out the molecular mechanisms underlying this unexpected BMAA effect, we studied the proteome of cyanobacterium Nostoc sp. PCC 7120 grown under BMAA treatment in nitrogen-replete medium. Experiments were performed in two experimental settings: (1) in control samples consisted of cells grown without the BMAA treatment and (2) the treated samples consisted of cells grown with addition of an aqueous solution of BMAA (20 µM). In total, 1567 different proteins of Nostoc sp. PCC 7120 were identified by LC-MS/MS spectrometry. Among them, 80 proteins belonging to different functional categories were chosen for further functional analysis and interpretation of obtained proteomic data. Here, we provide the evidence that a pleiotropic regulatory effect of BMAA on the proteome of cyanobacterium was largely different under conditions of nitrogen-excess compared to its effect under nitrogen starvation conditions (that was studied in our previous work). The most significant difference in proteome expression between the BMAA-treated and untreated samples under different growth conditions was detected in key regulatory protein PII (GlnB). BMAA downregulates protein PII in nitrogen-starved cells and upregulates this protein in nitrogen-replete conditions. PII protein is a key signal transduction protein and the change in its regulation leads to the change of many other regulatory proteins, including different transcriptional factors, enzymes and transporters. Complex changes in key metabolic and regulatory proteins (RbcL, RbcS, Rca, CmpA, GltS, NodM, thioredoxin 1, RpbD, ClpP, MinD, RecA, etc.), detected in this experimental study, could be a reason for the appearance of the “starvation” state in nitrogen-replete conditions in the presence of BMAA. In addition, 15 proteins identified in this study are encoded by genes, which are under the control of NtcA—a global transcriptional regulator—one of the main protein partners and transcriptional regulators of PII protein. Thereby, this proteomic study gives a possible explanation of cyanobacterium starvation under nitrogen-replete conditions and BMAA treatment. It allows to take a closer look at the regulation of cyanobacteria metabolism affected by this cyanotoxin. Full article

The oldest prokaryotic photoautotrophic organisms, cyanobacteria, produce many different metabolites. Among them is the water-soluble neurotoxic non-protein amino acid beta-N-methylamino-L-alanine (BMAA), whose biological functions in cyanobacterial metabolism are of fundamental scientific and practical interest. An early BMAA inhibitory effect on nitrogen fixation and heterocyst differentiation was shown in strains of diazotrophic cyanobacteria Nostoc sp. PCC 7120, Nostoc punctiforme PCC 73102 (ATCC 29133), and Nostoc sp. strain 8963 under conditions of nitrogen starvation. Herein, we present a comprehensive proteomic study of Nostoc (also called Anabaena) sp. PCC 7120 in the heterocyst formation stage affecting by BMAA treatment under nitrogen starvation conditions. BMAA disturbs proteins involved in nitrogen and carbon metabolic pathways, which are tightly co-regulated in cyanobacteria cells. The presented evidence shows that exogenous BMAA affects a key nitrogen regulatory protein, PII (GlnB), and some of its protein partners, as well as glutamyl-tRNA synthetase gltX and other proteins that are involved in protein synthesis, heterocyst differentiation, and nitrogen metabolism. By taking into account the important regulatory role of PII, it becomes clear that BMAA has a severe negative impact on the carbon and nitrogen metabolism of starving Nostoc sp. PCC 7120 cells. BMAA disturbs carbon fixation and the carbon dioxide concentrating mechanism, photosynthesis, and amino acid metabolism. Stress response proteins and DNA repair enzymes are upregulated in the presence of BMAA, clearly indicating severe intracellular stress. This is the first proteomic study of the effects of BMAA on diazotrophic starving cyanobacteria cells, allowing a deeper insight into the regulation of the intracellular metabolism of cyanobacteria by this non-protein amino acid. Full article

Small regulatory RNAs (sRNAs) are post-transcriptional regulators of bacterial gene expression. In cyanobacteria, the responses to nitrogen availability, that are mostly controlled at the transcriptional level by NtcA, involve also at least two small RNAs, namely NsiR4 (nitrogen stress-induced RNA 4) and NsrR1 (nitrogen stress-repressed RNA 1). Prediction of possible mRNA targets regulated by NsrR1 in Nostoc sp. PCC 7120 allowed, in addition to previously described nblA, the identification of all1871, a nitrogen-regulated gene encoding a protein of unknown function that we describe here as required for growth at the expense of atmospheric nitrogen (N₂). We show that transcription of all1871 is induced upon nitrogen step-down independently of NtcA. All1871 accumulation is repressed by NsrR1 and its expression is stronger in heterocysts, specialized cells devoted to N₂ fixation. We demonstrate specific interaction between NsrR1 and the 5′ untranslated region (UTR) of the all1871 mRNA, that leads to decreased expression of all1871. Because transcription of NsrR1 is partially repressed by NtcA, post-transcriptional regulation by NsrR1 would constitute an indirect way of NtcA-mediated regulation of all1871. Full article

Cyanobacteria produce a range of secondary metabolites, one being the neurotoxic non-protein amino acid β-N-methylamino-L-alanine (BMAA), proposed to be a causative agent of human neurodegeneration. As for most cyanotoxins, the function of BMAA in cyanobacteria is unknown. Here, we examined the effects of BMAA on the physiology of the filamentous nitrogen-fixing cyanobacterium Nostoc sp. PCC 7120. Our data show that exogenously applied BMAA rapidly inhibits nitrogenase activity (acetylene reduction assay), even at micromolar concentrations, and that the inhibition was considerably more severe than that induced by combined nitrogen sources and most other amino acids. BMAA also caused growth arrest and massive cellular glycogen accumulation, as observed by electron microscopy. With nitrogen fixation being a process highly sensitive to oxygen species we propose that the BMAA effects found here may be related to the production of reactive oxygen species, as reported for other organisms. Full article