Cyanobacterium Anabaena Sp Research Articles

Dependence of cyanobacterium growth and Mars-specific photobioreactor mass on total pressure, pN2 and pCO2

In situ resource utilization systems based on cyanobacteria could support the sustainability of crewed missions to Mars. However, their resource-efficiency will depend on the extent to which gases from the Martian atmosphere must be processed to support cyanobacterial growth. The main purpose of the present work is to help assess this extent. We therefore start with investigating the impact of changes in atmospheric conditions on the photoautotrophic, diazotrophic growth of the cyanobacterium Anabaena sp. PCC 7938. We show that lowering atmospheric pressure from 1 bar down to 80 hPa, without changing the partial pressures of metabolizable gases, does not reduce growth rates. We also provide equations, analogous to Monod’s, that describe the dependence of growth rates on the partial pressures of CO2 and N2. We then outline the relationships between atmospheric pressure and composition, the minimal mass of a photobioreactor’s outer walls (which is dependent on the inner-outer pressure difference), and growth rates. Relying on these relationships, we demonstrate that the structural mass of a photobioreactor can be decreased – without affecting cyanobacterial productivity – by reducing the inner gas pressure. We argue, however, that this reduction would be small next to the equivalent system mass of the cultivation system. A greater impact on resource-efficiency could come from the selection of atmospheric conditions which minimize gas processing requirements while adequately supporting cyanobacterial growth. The data and equations we provide can help identify these conditions.

Decrypting the effect of sulfate stress on the cyanobacterium Anabaena sp. PCC 7120 using physiological, proteome and transcript analysis in conjunction with bioinformatics

Sulfate limitation affects growth and various metabolic processes in cyanobacteria. However, there is little understanding of the underlying mechanism of these processes. Hence, the study investigated the effects of sulfate limitation in the cyanobacterium, Anabaena sp. PCC 7120. Results showed that sulfate stress affected photopigment contents in the test cyanobacterium delineating reduced growth. Proteome profile showed that differential expression profiles of different proteins involved in the various metabolic pathways suggested sulfur-dependent pathways were altered due to sulfate limitation. The most damaging effect was observed on the proteins belonging to the photosynthesis pathway, affirming the relationship between sulfur and carbon metabolism. Expression of proteins involved in the pathways such as nitrogen, protein synthesis, chaperone category, amino acid biosynthesis, and redox-responsive proteins were down-regulated. Moreover, forty-seven hypothetical proteins were also influenced by sulfate limitation. Molecular chaperones act as a physical barrier to prevent the aggregation of housekeeping proteins. Down expression of these proteins might affect the sustainability of the other proteins. Besides, transcript levels of the selected proteins were in concordance with physiological as well as proteome data. Additionally, template-based modeling and ab initio full atomic relaxation were performed via bioinformatic analysis through different tools to gain deeper insight into the structural and functional attributes of hypothetical proteins. Based on study, it could be concluded that mostly the proteins were downregulated which may lead to PCD in the cyanobacterium and also suggested that sulfur is a central hub and crucial for various sulfur-dependent pathways.

Analysis of fate determination of vegetative cells with reduced phycocyanin content in a multicellular cyanobacterium using Raman microscopy.

The one-dimensional multicellular cyanobacterium Anabaena sp. PCC 7120 exhibits two different cell types under nitrogen-deprived conditions. We found that the intensity of the Raman band at 1,629 cm -1 , which is associated with phycocyanin, was higher in undifferentiated cells (vegetative cells) than in differentiated cells (heterocysts). We observed cells whose band intensity at 1,629 cm -1 was statistically lower than that of vegetative cells, and named them "proheterocysts". We found that proheterocysts did not necessarily differentiate, and could divide or revert to being vegetative cells, as defined by having a higher band intensity at 1,629 cm -1 .

Two TonB-dependent outer membrane transporters involved in heme uptake in Anabaena sp. PCC 7120.

Low availability of micronutrients such as iron has enforced the evolution of uptake systems in all kingdoms of life. In Gram-negative bacteria, outer membrane, periplasmatic and plasma membrane localized proteins facilitate the uptake of iron-loaded chelators, which are energized by TonB proteins. The specificity of different uptake systems likely depends either on the endogenously produced siderophore or on the bioavailability of iron-chelator complexes in the environment. Hence, an uptake system for schizokinen produced by the model cyanobacterium Anabaena sp. PCC 7120 exists, while bioinformatics analysis suggests the existence of additional systems, likely for uptake of xenosiderophores. Consistently, proteins encoded by alr2153 (hutA1) and alr3242 (hutA2) are assigned as outer membrane heme transporters. Indeed, Anabaena sp. PCC 7120 can utilize external heme as an iron source. The addition of heme resulted in an induction of genes involved in heme degradation and chlorophyll a synthesis and in an increase of the chlorophyll a content. Moreover, iron starvation induced the expression of hutA1, while the addition of heme led to its repression. Remarkably, the addition of a high concentration of heme but not iron starvation resulted in hutA2 induction. Plasmid insertion mutants of both genes exhibited a reduced capacity to recover from iron starvation by heme addition, which indicates a dependence of heme uptake on functional HutA1 and HutA2 proteins. The structural model generated by bioinformatics methods is further in agreement with a role in heme uptake. Thus, we provide evidence that Anabaena sp. PCC 7120 uses a heme uptake system in parallel to other iron acquisition systems.

Exploring the potential of hydrogen sulfide in acquisition of aluminium stress tolerance in cyanobacterium Anabaena sp. PCC 7120

Exploring the potential of hydrogen sulfide in acquisition of aluminium stress tolerance in cyanobacterium Anabaena sp. PCC 7120

HetF defines a transition point from commitment to morphogenesis during heterocyst differentiation in the cyanobacterium Anabaena sp. PCC 7120.

The filamentous cyanobacterium Anabaena sp. PCC 7120 is able to form heterocysts for nitrogen fixation. Heterocyst differentiation is initiated by combined-nitrogen deprivation, followed by the commitment step during which the developmental process becomes irreversible. Mature heterocysts are terminally differentiated cells unable to divide, and cell division is required for heterocyst differentiation. Previously, we have shown that the HetF protease regulates cell division and heterocyst differentiation by cleaving PatU3, which is an inhibitor for both events. When hetF is required during the developmental program remains unknown. Here, by controlling the timing of hetF expression during heterocyst differentiation, we provide evidence that hetF is required just before the beginning of heterocyst morphogenesis. Consistent with this finding, transcriptome data show that most of the genes known to be involved in the early step (such as hetR and ntcA) or the commitment step (such as hetP and hetZ) of heterocyst development could be expressed in the ΔhetF mutant. In contrast, most of the genes involved in heterocyst morphogenesis and nitrogen fixation remain repressed in the mutant. These results indicated that in the absence of hetF, heterocyst differentiation is able to be initiated and proceeds to the stage just before heterocyst envelope formation.

Nitrogen and Redox Metabolism in Cyanobacterium Anabaena sp. PCC 7120 Exposed to Different Sulfate Regimes.

Sulfur is an important key nutrient required for the growth and development of cyanobacteria. Several reports showed the effect of sulfate limitation in unicellular and filamentous cyanobacteria, but such studies have not yet been reported in heterocytous cyanobacteria to ascribe the mechanisms of nitrogen and thiol metabolisms. Thus, the present work was carried out to appraise the impacts of sulfate limitation on nitrogen and thiol metabolisms in Anabaena sp. PCC 7120 by analyzing the contents as well as enzymes of nitrogen and thiol metabolisms. Cells of Anabaena sp. PCC 7120 were exposed to different regimes of sulfate, i.e., 300, 30, 3, and 0µM. Application of reduced concentration of sulfate showed negative impact on the cyanobacterium. Sulfate-limiting conditions reduces nitrogen-containing compounds in the cells of Anabaena. Additionally, reduced activities of nitrogen metabolic enzymes represented the role of sulfate in nitrogen metabolism. However, decreased activities of thiol metabolic enzymes indicated that sulfate-limited cyanobacterial cells have lower amount of glutathione and total thiol contents. Reduced accumulation of thiol components in thestressed cells indicated that sulfate-limited cells have lower ability to withstand stressful condition. Hence, Anabaena displays differential response to different concentrations of sulfate, and thus, stipulated that sulfur plays an important role in nitrogen and thiol metabolisms. To the best of our knowledge, this is the first report demonstrating the impact of sulfate stress on nitrogen and redox metabolisms in heterocytous cyanobacteria. This preliminary study provides a baseline idea that may help improve the production of paddy.

Tight association of CpcL with photosystem I in Anabaena sp. PCC 7120 grown under iron-deficient conditions

Phycobilisomes (PBSs), which are huge pigment-protein complexes displaying distinctive color variations, bind to photosystem cores for excitation-energy transfer. It is known that isolation of supercomplexes consisting of PBSs and photosystem I (PSI) or PBSs and photosystem II is challenging due to weak interactions between PBSs and the photosystem cores. In this study, we succeeded in purifying PSI-monomer-PBS and PSI-dimer-PBS supercomplexes from the cyanobacterium Anabaena sp. PCC 7120 grown under iron-deficient conditions by anion-exchange chromatography, followed by trehalose density gradient centrifugation. The absorption spectra of the two types of supercomplexes showed apparent bands originating from PBSs, and their fluorescence-emission spectra exhibited characteristic peaks of PBSs. Two-dimensional blue-native (BN)/SDS-PAGE of the two samples showed a band of CpcL, which is a linker protein of PBS, in addition to PsaA/B. Since interactions of PBSs with PSI are easily dissociated during BN-PAGE using thylakoids from this cyanobacterium grown under iron-replete conditions, it is suggested that iron deficiency for Anabaena induces tight association of CpcL with PSI, resulting in the formation of PSI-monomer-PBS and PSI-dimer-PBS supercomplexes. Based on these findings, we discuss interactions of PBSs with PSI in Anabaena.

Responses of Anabaena sp. PCC7120 to lindane: Physiological effects and differential expression of potential lin genes.

Lindane (γ-HCH) is an organochlorine pesticide that causes huge environmental concerns worldwide due to its recalcitrance and toxicity. The use of the cyanobacterium Anabaena sp. PCC 7120 in aquatic lindane bioremediation has been suggested but information relative to this process is scarce. In the present work, data relative to the growth, pigment composition, photosynthetic/respiration rate, and oxidative stress response of Anabaena sp. PCC 7120 in the presence of lindane at its solubility limit in water are shown. In addition, lindane degradation experiments revealed almost a total disappearance of lindane in the supernatants of Anabaena sp. PCC 7120 culture after 6 days of incubation. The diminishing in lindane concentration was in concordance with an increase in the levels of trichlorobenzene inside the cells. Furthermore, to identify potential orthologs of the linA, linB, linC, linD, linE, and linR genes from Sphingomonas paucimobilis B90A in Anabaena sp. PCC 7120, a whole genome screening was performed allowing the identification of five putative lin orthologs (all1353 and all0193 putative orthologs of linB, all3836 putative orthologs of linC, and all0352 and alr0353 putative orthologs of linE and linR, respectively)which could be involved in the lindane degradation pathway. Differential expression analysis of these genes in the presence of lindane revealed strong upregulation of one of the potential lin genes of Anabaena sp. PCC 7120.

Heterologous Production of the C33-C45 Polyketide Fragment of Anticancer Apratoxins in a Cyanobacterial Host.

A polyketide synthase subcluster of cytotoxic apratoxin A was isolated from a Moorena bouillonii environmental DNA library and engineered with a thioesterase II domain for heterologous expression in the filamentous cyanobacterium Anabaena sp. PCC7120. Further engineering with a rhamnose-inducible promoter led to the production of (2R,3R,5R,7R)-3,7-dihydroxy-2,5,8,8-tetramethylnonanoic acid, a stereogenically rich chiral building block that is important to the efficient synthesis of apratoxin analogues, representing the first synthetic biology attempt for this type of polyketide fragment.

The organization of the phycobilisome-photosystem I supercomplex depends on the ratio between two different phycobilisome linker proteins

The phycobilisome (PBS) is an antenna protein complex in cyanobacteria, Glaucocystophytes, and red algae. In the standard PBS, the rod-core PBS, the rods are connected to the core by the rod-core linker protein CpcG. The rod-core PBS transfers the light energy mainly to photosystem (PS) II and to a lesser extent to PSI. Cyanobacteria assemble another type of PBS, the CpcL-PBS, which consists of only one rod. This rod-type PBS is connected to the thylakoid membrane by the linker protein CpcL and is a PSI-specific antenna. In the filamentous heterocyst-forming cyanobacterium Anabaena (Nostoc) sp. PCC 7120, the CpcL-PBS forms a complex with the tetrameric PSI (PBS-PSI supercomplex). The CpcL-PBS and the rod part of the rod-core PBS are identical except for the linker proteins CpcL and CpcG. How cells control the accumulation of the two different types of PBS is unknown. Here, we analyzed two mutant strains which either lack the major rod-core linker CpcG4 or overexpress the rod-membrane linker CpcL. In both mutant strains, more and larger PBS-PSI supercomplexes accumulated compared to the wild type. Our results suggest that CpcL and CpcG4 compete for the same phycobiliprotein pool, and therefore the CpcL/CpcG4 ratio determines the levels of PBS-PSI supercomplexes. We propose that the CpcL-PBS and the rod-core PBS fulfill distinct functions in light harvesting.Graphical abstract

CvkR is a MerR-type transcriptional repressor of class 2 type V-K CRISPR-associated transposase systems

Certain CRISPR-Cas elements integrate into Tn7-like transposons, forming CRISPR-associated transposon (CAST) systems. How the activity of these systems is controlled in situ has remained largely unknown. Here we characterize the MerR-type transcriptional regulator Alr3614 that is encoded by one of the CAST (AnCAST) system genes in the genome of cyanobacterium Anabaena sp. PCC 7120. We identify a number of Alr3614 homologs across cyanobacteria and suggest naming these regulators CvkR for Cas V-K repressors. Alr3614/CvkR is translated from leaderless mRNA and represses the AnCAST core modules cas12k and tnsB directly, and indirectly the abundance of the tracr-CRISPR RNA. We identify a widely conserved CvkR binding motif 5’-AnnACATnATGTnnT-3’. Crystal structure of CvkR at 1.6 Å resolution reveals that it comprises distinct dimerization and potential effector-binding domains and that it assembles into a homodimer, representing a discrete structural subfamily of MerR regulators. CvkR repressors are at the core of a widely conserved regulatory mechanism that controls type V-K CAST systems.

Structure of a monomeric photosystem I core associated with iron-stress-induced-A proteins from Anabaena sp. PCC 7120

Iron-stress-induced-A proteins (IsiAs) are expressed in cyanobacteria under iron-deficient conditions. The cyanobacterium Anabaena sp. PCC 7120 has four isiA genes; however, their binding property and functional roles in PSI are still missing. We analyzed a cryo-electron microscopy structure of a PSI-IsiA supercomplex isolated from Anabaena grown under an iron-deficient condition. The PSI-IsiA structure contains six IsiA subunits associated with the PsaA side of a PSI core monomer. Three of the six IsiA subunits were identified as IsiA1 and IsiA2. The PSI-IsiA structure lacks a PsaL subunit; instead, a C-terminal domain of IsiA2 occupies the position of PsaL, which inhibits the oligomerization of PSI, leading to the formation of a PSI monomer. Furthermore, excitation-energy transfer from IsiAs to PSI appeared with a time constant of 55 ps. These findings provide insights into both the molecular assembly of the Anabaena IsiA family and the functional roles of IsiAs.

Cd-induced cytosolic proteome changes in the cyanobacterium Anabaena sp. PCC7120 are mediated by LexA as one of the regulatory proteins

LexA, a well-characterized transcriptional repressor of SOS genes in heterotrophic bacteria, has been shown to regulate diverse genes in cyanobacteria. An earlier study showed that LexA overexpression in a cyanobacterium, Anabaena sp. PCC7120 reduces its tolerance to Cd stress. This was later shown to be due to modulation of photosynthetic redox poising by LexA under Cd stress. However, due to the global regulatory nature of LexA and the prior prediction of AnLexA-box in a few heavy metal-responsive genes, we speculated that LexA has a broad role in Cd tolerance, with regulation over a variety of Cd stress-responsive genes in addition to photosynthetic genes. Thus, to further expand the knowledge on the regulatory role of LexA in Cd stress tolerance, a cytosolic proteome profiling of Anabaena constitutively overexpressing LexA upon Cd stress was performed. The proteomic study revealed 25 differentially accumulated proteins (DAPs) in response to the combined effect of LexA overexpression and Cd stress, and the other 11 DAPs exclusively in response to either LexA overexpression or Cd stress. The 36 identified proteins were related with a variety of functions, including photosynthesis, C-metabolism, antioxidants, protein turnover, post-transcriptional modifications, and a few unknown and hypothetical proteins. The regulation of LexA on corresponding genes, and six previously reported Cd efflux transporters, was further validated by the presence of AnLexA-boxes, transcript, and/or promoter analyses. In a nutshell, this study identifies the regulation of Anabaena LexA on several Cd stress-responsive genes of various functions, hence expanding the regulatory role of LexA under Cd stress.

Gamma (γ)-radiation stress response of the cyanobacterium Anabaena sp. PCC7120: Regulatory role of LexA and photophysiological changes

High radioresistance of the cyanobacterium, Anabaena sp. PCC7120 has been attributed to efficient DNA repair, protein recycling, and oxidative stress management. However, the regulatory network involved in these batteries of responses remains unexplored. In the present study, the role of a global regulator, LexA in modulating gamma (γ)-radiation stress response of Anabaena was investigated. Comparison of the cytosolic proteome profiles upon γ-radiation in recombinant Anabaena strains, AnpAM (vector-control) and AnlexA+ (LexA-overexpressing), revealed 41 differentially accumulated proteins, corresponding to 29 distinct proteins. LexA was found to be involved in the regulation of 27 of the corresponding genes based on the presence of AnLexA-Box, EMSA, and/or qRT-PCR studies. The majority of the regulated genes were found to be involved in C-assimilation either through photosynthesis or C-catabolism and oxidative stress alleviation. Photosynthesis, measured in terms of PSII photophysiological parameters and thylakoid membrane proteome was found to be affected by γ-radiation in both AnpAM and AnlexA+ cells, with LexA affecting them even under control growth conditions. Thus, LexA functioned as one of the transcriptional regulators involved in modulating γ-radiation stress response in Anabaena. This study could pave the way for a deeper understanding of the regulation of γ-radiation-responsive genes in cyanobacteria at large.

Flv3A facilitates O2 photoreduction and affects H2 photoproduction independently of Flv1A in diazotrophic Anabaena filaments.

The model heterocyst-forming filamentous cyanobacterium Anabaena sp. PCC 7120 (Anabaena) is a typical example of a multicellular organism capable of simultaneously performing oxygenic photosynthesis in vegetative cells and O2 -sensitive N2 -fixation inside heterocysts. The flavodiiron proteins have been shown to participate in photoprotection of photosynthesis by driving excess electrons to O2 (a Mehler-like reaction). Here, we performed a phenotypic and biophysical characterization of Anabaena mutants impaired in vegetative-specific Flv1A and Flv3A in order to address their physiological relevance in the bioenergetic processes occurring in diazotrophic Anabaena under variable CO2 conditions. We demonstrate that both Flv1A and Flv3A are required for proper induction of the Mehler-like reaction upon a sudden increase in light intensity, which is likely important for the activation of carbon-concentrating mechanisms and CO2 fixation. Under ambient CO2 diazotrophic conditions, Flv3A is responsible for moderate O2 photoreduction, independently of Flv1A, but only in the presence of Flv2 and Flv4. Strikingly, the lack of Flv3A resulted in strong downregulation of the heterocyst-specific uptake hydrogenase, which led to enhanced H2 photoproduction under both oxic and micro-oxic conditions. These results reveal a novel regulatory network between the Mehler-like reaction and the diazotrophic metabolism, which is of great interest for future biotechnological applications.

The Role of MreB, MreC and MreD in the Morphology of the Diazotrophic Filament of Anabaena sp. PCC 7120.

The cyanobacterium Anabaena sp. PCC 7120 forms filaments of communicating cells. Under conditions of nitrogen scarcity, some cells differentiate into heterocysts, allowing the oxygen-sensitive N2-reduction system to be expressed and operated in oxic environments. The key to diazotrophic growth is the exchange of molecules with nutritional and signaling functions between the two types of cells of the filament. During heterocyst differentiation, the peptidoglycan sacculus grows to allow cell enlargement, and the intercellular septa are rebuilt to narrow the contact surface with neighboring cells and to hold specific transport systems, including the septal junction complexes for intercellular molecular transfer, which traverse the periplasm between heterocysts and neighboring vegetative cells through peptidoglycan nanopores. Here we have followed the spatiotemporal pattern of peptidoglycan incorporation during heterocyst differentiation by Van-FL labeling and the localization and role of proteins MreB, MreC and MreD. We observed strong transitory incorporation of peptidoglycan in the periphery and septa of proheterocysts and a maintained focal activity in the center of mature septa. During differentiation, MreB, MreC and MreD localized throughout the cell periphery and at the cell poles. In mreB, mreC or mreD mutants, instances of strongly increased peripheral and septal peptidoglycan incorporation were detected, as were also heterocysts with aberrant polar morphology, even producing filament breakage, frequently lacking the septal protein SepJ. These results suggest a role of Mre proteins in the regulation of peptidoglycan growth and the formation of the heterocyst neck during differentiation, as well as in the maintenance of polar structures for intercellular communication in the mature heterocyst. Finally, as previously observed in filaments growing with combined nitrogen, in the vegetative cells of diazotrophic filaments, the lack of MreB, MreC or MreD led to altered localization of septal peptidoglycan-growth bands reproducing an altered localization of FtsZ and ZipN rings during cell division.

Identification and toxicity towards aquatic primary producers of the smallest fractions released from hydrolytic degradation of polycaprolactone microplastics.

Bioplastics are thought as a safe substitute of non-biodegradable polymers. However, once released in the environment, biodegradation may be very slow, and they also suffer abiotic fragmentation processes, which may give rise to different fractions of polymer sizes. We present novel data on abiotic hydrolytic degradation of polycaprolactone (PCL), tracking the presence of by-products during 132 days by combining different physicochemical techniques. During the study a considerable amount of two small size plastic fractions were found (up to∼6mg of PCL by-product/g of PCL beads after 132 days of degradation); and classified as submicron-plastics (sMPs) from 1μm to 100nm and nanoplastics (NPs, <100nm) as well as oligomers. The potential toxicity of the smallest fractions, PCL by-products < 100nm (PCL-NPs+PCL oligomers) and the PCL oligomers single fraction, was tested on two ecologically relevant aquatic primary producers: the heterocystous filamentous nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120, and the unicellular cyanobacterium Synechococcus sp. PCC 7942. Upon exposure to both, single and combined fractions, Reactive Oxygen Species (ROS) overproduction, intracellular pH and metabolic activity alterations were observed in both organisms, whilst membrane potential and morphological damages were only observed upon PCL-NPs+PCL oligomers exposure. Notably both PCL by-products fractions inhibited nitrogen fixation in Anabaena, which may be clearly detrimental for the aquatic trophic chain. As conclusion, fragmentation of bioplastics may render a continuous production of secondary nanoplastics as well as oligomers that might be toxic to the surrounding biota; both PCL-NPs and PCL oligomers, but largely the nanoparticulate fraction, were harmful for the two aquatic primary producers. Efforts should be made to thoroughly understand the fragmentation of bioplastics and the toxicity of the smallest fractions resulting from that degradation.

The occurrence of the cis/trans geometric isomerism of myxoxanthophyll and 4-ketomyxoxanthophyll in the cyanobacterium Anabaena sp. PCC7120.

The cyanobacteria of the genus Anabaena have recently been recognized as a potential source of secondary metabolites of pharmacological and biotechnological importance. In particular, myxoxanthophylls - specific carotenoid glycosides that accumulate in cyanobacterial cells, are attracting increasing interest. Anabaena (Nostoc) sp. PCC7120, a filamentous, mesophilic, nitrogen-fixing cyanobacterium, is a model organism used in biochemical and genetic studies. The carotenoid pool of Anabaena sp. PCC7120 consists of five main species of pigments, namely β-carotene, echinenone, canthaxanthin and two derivatives of myxoxanthophyll: myxoxanthophyll ((3R,2'S)-myxol 2'-fucoside) and 4-ketomyxoxanthophyll ((3S,2'S)-4-ketomyxol 2'-fucoside). Recent findings show that the carotenoid biosynthesis pathway functions in Anabaena sp. PCC7120 cells are affected by environmental factors. Specifically, the balance between β-carotene and ketocarotenoids alters according to the temperature conditions. In this study, a new method, based on single-step liquid adsorption chromatography was developed and applied to separate a fraction containing myxoxanthophyll and 4-ketomyxoxanthophyll from Anabaena sp. PCC7120 cells. It was found that this method allowed a high purity fraction of carotenoid glycosides to be obtained from pigment pools as extracted from cyanobacterial cells. The subsequent analysis using the methods HPLC and LC/MS demonstrated that this fraction consists of a mixture of compounds with different retention times. On the basis of their fragmentation spectra and optical properties, these compounds were identified as geometrical isomers of myxoxanthophyll and 4-ketomyxoxanthophyll, including the dominant all-trans forms and less abundant cis forms. Proposals regarding the structures of myxoxanthophyll isomers are made.

A proteolytic pathway coordinates cell division and heterocyst differentiation in the cyanobacterium Anabaena sp. PCC 7120

The filamentous, multicellular cyanobacterium Anabaena sp. PCC 7120 (Anabaena) is a prokaryotic model for the study of cell differentiation and cell-cell interactions. Upon combined-nitrogen deprivation, Anabaena forms a particular cell type, heterocyst, for aerobic nitrogen fixation. Heterocysts are semiregularly spaced among vegetative cells. Heterocyst differentiation is coupled to cell division, but the underlying mechanism remains unclear. This mechanism could be mediated by the putative protease HetF, which is a divisome component and is necessary for heterocyst differentiation. In this study, by suppressor screening, we identified PatU3, as a negative regulator acting downstream of HetF for cell division and heterocyst development. The inactivation of patU3 restored the capacity of cell division and heterocyst differentiation in the ΔhetF mutant, and overexpression of patU3 inhibited both processes in the wild-type background. We demonstrated that PatU3 was a specific substrate of the protease activity of HetF. Consequently, PatU3 accumulated in the hetF-deficient mutant, which was responsible for the resultant mutant phenotype. The cleavage site of PatU3 by HetF was mapped after the Arg117 residue, whose mutation made PatU3 resistant to HetF processing, and mimicked the effect of hetF deletion. Our results provided evidence that HetF regulated cell division and heterocyst differentiation by controlling the inhibitory effects of PatU3. This proteolytic pathway constituted a mechanism for the coordination between cell division and differentiation in a prokaryotic model used for studies on developmental biology and multicellularity.