Comparative genomics reveals insights into cyanobacterial evolution and habitat adaptation

Cyanobacteria are photosynthetic prokaryotes that inhabit diverse aquatic and terrestrial environments. However, the evolutionary mechanisms involved in the cyanobacterial habitat adaptation remain poorly understood. Here, based on phylogenetic and comparative genomic analyses of 650 cyanobacterial genomes, we investigated the genetic basis of cyanobacterial habitat adaptation (marine, freshwater, and terrestrial). We show: (1) the expansion of gene families is a common strategy whereby terrestrial cyanobacteria cope with fluctuating environments, whereas the genomes of many marine strains have undergone contraction to adapt to nutrient-poor conditions. (2) Hundreds of genes are strongly associated with specific habitats. Genes that are differentially abundant in genomes of marine, freshwater, and terrestrial cyanobacteria were found to be involved in light sensing and absorption, chemotaxis, nutrient transporters, responses to osmotic stress, etc., indicating the importance of these genes in the survival and adaptation of organisms in specific habitats. (3) A substantial fraction of genes that facilitate the adaptation of Cyanobacteria to specific habitats are contributed by horizontal gene transfer, and such genetic exchanges are more frequent in terrestrial cyanobacteria. Collectively, our results further our understandings of the adaptations of Cyanobacteria to different environments, highlighting the importance of ecological constraints imposed by the environment in shaping the evolution of Cyanobacteria.Subject terms: Phylogenetics, Microbial ecology     

Genome-wide identification of major protein families of cyanobacteria and genomic insight into the circadian rhythm

Cyanobacteria are among the most abundant organisms present on earth and are considered to be one of the oldest known clades. Cyanobacteria are oxygenic photosynthetic bacteria and are well known as promising renewable sources of energy; therefore, it is important to understand aspects of their genomes in detail across species. Advances in sequencing technology and the availability of several cyanobacterial genomes have provided an excellent opportunity to understand the diversity and evolution of the cyanobacterial genome. Here, we compared the genomes of 62 different phototrophic cyanobacteria. Evaluation of genetic diversity of all the cyanobacteria species studied revealed that evolution from their common ancestors was polyphyletic. In addition, the genomes were very diverse and varied among species, and significant genomic diversity was observed at the species and strain level. Overall, we identified 56 different protein families of cyanobacteria species/strains and found that they varied significantly among strains of a species. The circadian clock proteins KaiA, KaiB and KaiC (KaiABC complex proteins) of cyanobacteria were found to be present and consistent in the majority of cyanobacterial species while absent in a few others. Evolutionary analysis of the KaiABC protein complex showed that the KaiA protein has a high frequency of polymorphism, and multiple alleles were found to be present at high frequency. These results demonstrated that evolution of phosphorylation events occurred via KaiA in the KaiABC complex. Furthermore, multiple sequence alignment showed that KaiA, KaiB and KaiC proteins are highly conserved in nature. Our results provide direct information regarding the presence of different protein or protein families in cyanobacteria. The information presented here will serve as a starting point to explore the genetic diversity of cyanobacteria with the potential to play important roles in biotechnological applications.     

Genome mining reveals high incidence of putative lipopeptide biosynthesis NRPS/PKS clusters containing fatty acyl‐AMP ligase genes in biofilm‐forming cyanobacteria

Cyanobacterial lipopeptides have antimicrobial and antifungal bioactivities with potential for use in pharmaceutical research. However, due to their hemolytic activity and cytotoxic effects on human cells, they may pose a health issue if produced in substantial amounts in the environment. In bacteria, lipopeptides can be synthesized via several well‐evidenced mechanisms. In one of them, fatty acyl‐AMP ligase (FAAL) initiates biosynthesis by activation of a fatty acyl residue. We have performed a bioinformatic survey of the cyanobacterial genomic information available in the public databases for the presence of FAAL‐containing non‐ribosomal peptide synthetase/polyketide synthetase (NRPS/PKS) biosynthetic clusters, as a genetic basis for lipopeptide biosynthesis. We have identified 79 FAAL genes associated with various NRPS/PKS clusters in 16% of 376 cyanobacterial genomic assemblies available, suggesting that FAAL is frequently incorporated in NRPS/PKS biosynthetases. FAAL was present either as a stand‐alone protein or fused either to NRPS or PKS. Such clusters were more frequent in derived phylogenetic lineages with larger genome sizes, which is consistent with the general pattern of NRPS/PKS pathways distribution. The putative lipopeptide clusters were more frequently found in genomes of cyanobacteria that live attached to surfaces and are capable of forming microbial biofilms. While lipopeptides are known in other bacterial groups to play a role in biofilm formation, motility, and colony expansion, their functions in cyanobacterial biofilms need to be tested experimentally. According to our data, benthic and terrestrial cyanobacteria should be the focus of a search for novel candidates for lipopeptide drug synthesis and the monitoring of toxic lipopeptide production.     

Gene pool transmission of multidrug resistance among Campylobacter from livestock,sewage and human disease

The use of antimicrobials in human and veterinary medicine has coincided with a rise in antimicrobial resistance (AMR) in the food-borne pathogens Campylobacter jejuni and Campylobacter coli. Faecal contamination from the main reservoir hosts (livestock, especially poultry) is the principal route of human infection but little is known about the spread of AMR among source and sink populations. In particular, questions remain about how Campylobacter resistomes interact between species and hosts, and the potential role of sewage as a conduit for the spread of AMR. Here, we investigate the genomic variation associated with AMR in 168 C. jejuni and 92 C. coli strains isolated from humans, livestock and urban effluents in Spain. AMR was tested in vitro and isolate genomes were sequenced and screened for putative AMR genes and alleles. Genes associated with resistance to multiple drug classes were observed in both species and were commonly present in multidrug-resistant genomic islands (GIs), often located on plasmids or mobile elements. In many cases, these loci had alleles that were shared among C. jejuni and C. coli consistent with horizontal transfer. Our results suggest that specific antibiotic resistance genes have spread among Campylobacter isolated from humans, animals and the environment.     

Heme-copper oxidases and their electron donors in cyanobacterial respiratory electron transport

Cyanobacteria are the paradigmatic organisms of oxygenic (plant-type) photosynthesis and aerobic respiration. Since there is still an amazing lack of knowledge on the role and mechanism of their respiratory electron transport, we have critically analyzed all fully or partially sequenced genomes for heme-copper oxidases and their (putative) electron donors cytochrome c(6), plastocyanin, and cytochrome c(M). Well-known structure-function relationships of the two branches of heme-copper oxidases, namely cytochrome c (aa(3)-type) oxidase (COX) and quinol (bo-type) oxidase (QOX), formed the base for a critical inspection of genes and ORFs found in cyanobacterial genomes. It is demonstrated that at least one operon encoding subunits I-III of COX is found in all cyanobacteria, whereas many non-N(2)-fixing species lack QOX. Sequence analysis suggests that both cyanobacterial terminal oxidases should be capable of both the four-electron reduction of dioxygen and proton pumping. All diazotrophic organisms have at least one operon that encodes QOX. In addition, the highly refined specialization in heterocyst forming Nostocales is reflected by the presence of two paralogs encoding COX. The majority of cyanobacterial genomes contain one gene or ORF for plastocyanin and cytochrome c(M), whereas 1-4 paralogs for cytochrome c(6) were found. These findings are discussed with respect to published data about the role of respiration in wild-type and mutated cyanobacterial strains in normal metabolism, stress adaptation, and nitrogen fixation. A model of the branched electron-transport pathways downstream of plastoquinol in cyanobacteria is presented.     

Genes of cyanobacterial origin in plant nuclear genomes point to a heterocyst-forming plastid ancestor

Plastids are descended from a cyanobacterial symbiosis which occurred over 1.2 billion years ago. During the course of endosymbiosis, most genes were lost from the cyanobacterium's genome and many were relocated to the host nucleus through endosymbiotic gene transfer (EGT). The issue of how many genes were acquired through EGT in different plant lineages is unresolved. Here, we report the genome-wide frequency of gene acquisitions from cyanobacteria in 4 photosynthetic eukaryotes--Arabidopsis, rice, Chlamydomonas, and the red alga Cyanidioschyzon--by comparison of the 83,138 proteins encoded in their genomes with 851,607 proteins encoded in 9 sequenced cyanobacterial genomes, 215 other reference prokaryotic genomes, and 13 reference eukaryotic genomes. The analyses entail 11,569 phylogenies inferred with both maximum likelihood and Neighbor-Joining approaches. Because each phylogenetic result is dependent not only upon the reconstruction method but also upon the site patterns in the underlying alignment, we investigated how the reliability of site pattern generation via alignment affects our results: if the site patterns in an alignment differ depending upon the order in which amino acids are introduced into multiple sequence alignment--N- to C-terminal versus C- to N-terminal--then the phylogenetic result is likely to be artifactual. Excluding unreliable alignments by this means, we obtain a conservative estimate, wherein about 14% of the proteins examined in each plant genome indicate a cyanobacterial origin for the corresponding nuclear gene, with higher proportions (17-25%) observed among the more reliable alignments. The identification of cyanobacterial genes in plant genomes affords access to an important question: from which type of cyanobacterium did the ancestor of plastids arise? Among the 9 cyanobacterial genomes sampled, Nostoc sp. PCC7120 and Anabaena variabilis ATCC29143 were found to harbor collections of genes which are-in terms of presence/absence and sequence similarity-more like those possessed by the plastid ancestor than those of the other 7 cyanobacterial genomes sampled here. This suggests that the ancestor of plastids might have been an organism more similar to filamentous, heterocyst-forming (nitrogen-fixing) representatives of section IV recognized in Stanier's cyanobacterial classification. Members of section IV are very common partners in contemporary symbiotic associations involving endosymbiotic cyanobacteria, which generally provide nitrogen to their host, consistent with suggestions that fixed nitrogen supplied by the endosymbiont might have played an important role during the origin of plastids.     

Simultaneous quantification of cyanobacteria and Microcystis spp. using real-time PCR

In order to develop a protocol to quantify cyanobacteria and Microcystis simultaneously, the primers and probe were designed from the conserved regions of 16S rRNA gene sequences of cyanobacteria and Microcystis, respectively. Probe match analysis of the Ribosomal Database Project showed that the primers matched with over 97% of cyanobacterial 16S rRNA genes, indicating these can be used to amplify cyanobacteria specifically. The TaqMan probe, which is located between two primers, matched with 98.2% of sequences in genus GpXI, in which most Microcystis strains are included. The numbers of cyanobacterial genes were estimated with the emission of SYBR Green from the amplicons with two primers, whereas those of Microcystis spp. were measured from the fluorescence of CAL Fluor Gold 540 emitted by exonuclease activity of Taq DNA polymerase in amplification. It is expected that this method enhances the accuracy and reduces the time to count cyanobacteria and potential toxigenic Microcystis spp. in aquatic environmental samples.     

Cyanobactins—ribosomal cyclic peptides produced by cyanobacteria

Cyanobactins are small cyclic peptides that are produced by a diverse selection of cyanobacteria living in symbioses as well as terrestrial, marine, or freshwater environments. They include compounds with antimalarial, antitumor, and multidrug reversing activities and potential as pharmaceutical leads. Cyanobactins are produced through the proteolytic cleavage and cyclization of precursor peptides coupled with further posttranslational modifications such as heterocyclization, oxidation, or prenylation of amino acids. Cyanobactin gene clusters encode two proteases which cleave and cyclisize the precursor peptide as well as proteins participating in posttranslational modifications. The bioinformatic mining of cyanobacterial genomes has led to the discovery of novel cyanobactins. Heterologous expression of these gene clusters provided insights into the role of the genes participating in the biosynthesis of cyanobactins and facilitated the rational design of novel peptides. Enzymes participating in the biosynthesis of cyanobactins may prove useful as catalysts for producing novel cyclic peptides in the future. The recent discovery of the cyanobactin biosynthetic pathway in cyanobacteria extends our knowledge of their potential as producers of interesting metabolites.     

Distribution and diversity of natural product genes in marine and freshwater cyanobacterial cultures and genomes

Natural products are a functionally diverse class of biochemically synthesized compounds, which include antibiotics, toxins, and siderophores. In this paper, we describe both the detection of natural product activities and the sequence identification of gene fragments from two molecular systems that have previously been implicated in natural product production, i.e., nonribosomal peptide synthetases (NRPSs) and modular polyketide synthases (PKSs), in diverse marine and freshwater cyanobacterial cultures. Using degenerate PCR and the sequencing of cloned products, we show that NRPSs and PKSs are common among the cyanobacteria tested. Our molecular data, when combined with genomic searches of finished and progressing cyanobacterial genomes, demonstrate that not all cyanobacteria contain NRPS and PKS genes and that the filamentous and heterocystous cyanobacteria are the richest sources of these genes and the most likely sources of novel natural products within the phylum. In addition to validating the use of degenerate primers for the identification of PKS and NRPS genes in cyanobacteria, this study also defines numerous gene fragments that will be useful as probes for future studies of the synthesis of natural products in cyanobacteria. Phylogenetic analyses of the cyanobacterial NRPS and PKS fragments sequenced in this study, as well as those from the cyanobacterial genome projects, demonstrate that there is remarkable diversity and likely novelty of these genes within the cyanobacteria. These results underscore the potential variety of novel products being produced by these ubiquitous organisms.     

Coastal Synechococcus metagenome reveals major roles for horizontal gene transfer and plasmids in population diversity

The extent to which cultured strains represent the genetic diversity of a population of microorganisms is poorly understood. Because they do not require culturing, metagenomic approaches have the potential to reveal the genetic diversity of the microbes actually present in an environment. From coastal California seawater, a complex and diverse environment, the marine cyanobacteria of the genus Synechococcus were enriched by flow cytometry-based sorting and the population metagenome was analysed with 454 sequencing technology. The sequence data were compared with model Synechococcus genomes, including those of two coastal strains, one isolated from the same and one from a very similar environment. The natural population metagenome had high sequence identity to most genes from the coastal model strains but diverged greatly from these genomes in multiple regions of atypical trinucleotide content that encoded diverse functions. These results can be explained by extensive horizontal gene transfer presumably with large differences in horizontally transferred genetic material between different strains. Some assembled contigs showed the presence of novel open reading frames not found in the model genomes, but these could not yet be unambiguously assigned to a Synechococcus clade. At least three distinct mobile DNA elements (plasmids) not found in model strain genomes were detected in the assembled contigs, suggesting for the first time their likely importance in marine cyanobacterial populations and possible role in horizontal gene transfer.     

Analysis of the hli gene family in marine and freshwater cyanobacteria

Certain cyanobacteria thrive in natural habitats in which light intensities can reach 2000 micromol photon m(-2) s(-1) and nutrient levels are extremely low. Recently, a family of genes designated hli was demonstrated to be important for survival of cyanobacteria during exposure to high light. In this study we have identified members of the hli gene family in seven cyanobacterial genomes, including those of a marine cyanobacterium adapted to high-light growth in surface waters of the open ocean (Prochlorococcus sp. strain Med4), three marine cyanobacteria adapted to growth in moderate- or low-light (Prochlorococcus sp. strain MIT9313, Prochlorococcus marinus SS120, and Synechococcus WH8102), and three freshwater strains (the unicellular Synechocystis sp. strain PCC6803 and the filamentous species Nostoc punctiforme strain ATCC29133 and Anabaena sp. [Nostoc] strain PCC7120). The high-light-adapted Prochlorococcus Med4 has the smallest genome (1.7 Mb), yet it has more than twice as many hli genes as any of the other six cyanobacterial species, some of which appear to have arisen from recent duplication events. Based on cluster analysis, some groups of hli genes appear to be specific to either marine or freshwater cyanobacteria. This information is discussed with respect to the role of hli genes in the acclimation of cyanobacteria to high light, and the possible relationships among members of this diverse gene family.     

Cyanobacteria in Antarctica: ecology, physiology and cold adaptation.

Cyanobacterial species composition of fresh water and terrestrial ecosystems and chemical environment of water in Schirmacher Oasis in Continental Antarctica was investigated. Over 35 species of cyanobacteria were recorded. Diazotrophic species both heterocystous and unicellular contributed more than half to the count except in lake ecosystem. The species composition varied among the fresh water as well as terrestrial ecosystems. The physico-chemical analyses of water revealed its poor nurient content which might have supported the growth of diazotrophic cyanobacteria in an Antarctic environment. Among the cyanobacteria Oscillatoria, Phormidium and Nostoc commune were the dominant flora in most of the habitats. The physiological characteristics of isolated cyanobacteria strains indicated that N2-fixation, nitrate uptake, nitrate-reduction, ammonium-uptake, GS-transferase activity and photosynthesis was unaffected at low temperature (5 degrees C) which indicated low temperature adaptation for Antarctic cyanobacteria. This phenomenon was not evident in different strains of tropical origin. The temperature optima for N2-fixation for the different Antarctic cyanobacterial strains was in the range of 15-25 degrees C, nearly 10 degrees C lower than their respective reference strains of tropical origin. Similar results were obtained for cyanobacteria-moss association. The low endergonic activation energy exhibited by the above metabolic activities supported the view that cyanobacteria were adapted to Antarctic ecosystem.     

Diversity and expression of cyanobacterial hupS genes in pure cultures and in a nitrogen-limited phototrophic biofilm

A PCR was developed for conserved regions within the cyanobacterial small subunit uptake hydrogenase (hupS) gene family. These primers were used to PCR amplify partial hupS sequences from 15 cyanobacterial strains. HupS clone libraries were constructed from PCR-amplified genomic DNA and reverse-transcribed mRNA extracted from phototrophic biofilms cultivated under nitrate-limiting conditions. Partial hupS gene sequences derived from cyanobacteria, some of which were not previously known to contain hup genes were used for phylogenetic analysis. Phylogenetic trees constructed with partial hupS genes were congruent with those based on 16S rRNA genes, indicating that hupS sequences can be used to identify cyanobacteria expressing hup. Sequences from heterocystous and nonheterocystous cyanobacteria formed two separate clusters. Analysis of clone library data showed a discrepancy between the presence and the activity of cyanobacterial hupS genes in phototrophic biofilms. The results showed that the hupS gene can be used to characterize the diversity of natural populations of diazotrophic cyanobacteria, and to characterize gene expression patterns of individual species and strains.     

Selection of cyanobacteria isolated from mosquito breeding sites as a potential food source for mosquito larvae.

One way to increase the persistence of larvicidal toxins in mosquito breeding sites is to clone the corresponding genes in microorganisms, such as cyanobacteria, which could serve as a source of food for the larvae. We isolated and cultured 10 strains of cyanobacteria from three mosquito breeding sites along the French Mediterranean coast. Most of the strains were tolerant to a relatively wide range of salt concentrations, and all of them were totally or partially resistant to at least four of the five biological or chemical larvicides used in the local mosquito control program. Six unicellular strains from these habitats and Synechococcus strain PCC 7942, a strain maintained for more than 10 years under laboratory conditions, were assessed for ingestion and digestion by larvae Culex pipiens and Anopheles gambiae mosquitoes. The numbers of cells ingested and digested were dependent on the cyanobacterial strain and varied with the mosquito species. Three of the new isolates, Synechococcus strain PCC 8905 and Synechocystis strains PCC 8906 and PCC 8912, were ingested and digested rapidly by larvae of both mosquito species. Since these strains are also tolerant to larvicides and relatively resistant to elevated salt concentrations, they meet the basic requirements for potential recipients of bacterial genes that encode endotoxins.     

Selection of cyanobacteria isolated from mosquito breeding sites as a potential food source for mosquito larvae.

One way to increase the persistence of larvicidal toxins in mosquito breeding sites is to clone the corresponding genes in microorganisms, such as cyanobacteria, which could serve as a source of food for the larvae. We isolated and cultured 10 strains of cyanobacteria from three mosquito breeding sites along the French Mediterranean coast. Most of the strains were tolerant to a relatively wide range of salt concentrations, and all of them were totally or partially resistant to at least four of the five biological or chemical larvicides used in the local mosquito control program. Six unicellular strains from these habitats and Synechococcus strain PCC 7942, a strain maintained for more than 10 years under laboratory conditions, were assessed for ingestion and digestion by larvae Culex pipiens and Anopheles gambiae mosquitoes. The numbers of cells ingested and digested were dependent on the cyanobacterial strain and varied with the mosquito species. Three of the new isolates, Synechococcus strain PCC 8905 and Synechocystis strains PCC 8906 and PCC 8912, were ingested and digested rapidly by larvae of both mosquito species. Since these strains are also tolerant to larvicides and relatively resistant to elevated salt concentrations, they meet the basic requirements for potential recipients of bacterial genes that encode endotoxins.     

Evolution of enzymes involved in the photorespiratory 2-phosphoglycolate cycle from cyanobacteria via algae toward plants

The photorespiratory pathway was shown to be essential for organisms performing oxygenic photosynthesis, cyanobacteria, algae, and plants, in the present day O(2)-containing atmosphere. The identification of a plant-like 2-phosphoglycolate cycle in cyanobacteria indicated that not only genes of oxygenic photosynthesis but also genes encoding photorespiratory enzymes were endosymbiotically conveyed from ancient cyanobacteria to eukaryotic oxygenic phototrophs. Here, we investigated the origin of the photorespiratory pathway in photosynthetic eukaryotes by phylogenetic analysis. We found that a mixture of photorespiratory enzymes of either cyanobacterial or α-proteobacterial origin is present in algae and higher plants. Three enzymes in eukaryotic phototrophs clustered closely with cyanobacterial homologs: glycolate oxidase, glycerate kinase, and hydroxypyruvate reductase. On the other hand, the mitochondrial enzymes of the photorespiratory cycle in algae and plants, glycine decarboxylase subunits and serine hydroxymethyltransferase, evolved from proteobacteria. Other than most genes for proteins of the photosynthetic machinery, nearly all enzymes involved in the 2-phosphogylcolate metabolism coexist in the genomes of cyanobacteria and heterotrophic bacteria.     

Dryland biological soil crust cyanobacteria show unexpected decreases in abundance under long‐term elevated CO2

Biological soil crusts (biocrusts) cover soil surfaces in many drylands globally. The impacts of 10 years of elevated atmospheric CO₂ on the cyanobacteria in biocrusts of an arid shrubland were examined at a large manipulated experiment in Nevada, USA. Cyanobacteria‐specific quantitative PCR surveys of cyanobacteria small‐subunit (SSU) rRNA genes suggested a reduction in biocrust cyanobacterial biomass in the elevated CO₂ treatment relative to the ambient controls. Additionally, SSU rRNA gene libraries and shotgun metagenomes showed reduced representation of cyanobacteria in the total microbial community. Taxonomic composition of the cyanobacteria was similar under ambient and elevated CO₂ conditions, indicating the decline was manifest across multiple cyanobacterial lineages. Recruitment of cyanobacteria sequences from replicate shotgun metagenomes to cyanobacterial genomes representing major biocrust orders also suggested decreased abundance of cyanobacteria sequences across the majority of genomes tested. Functional assignment of cyanobacteria‐related shotgun metagenome sequences indicated that four subsystem categories, three related to oxidative stress, were differentially abundant in relation to the elevated CO₂ treatment. Taken together, these results suggest that elevated CO₂ affected a generalized decrease in cyanobacteria in the biocrusts and may have favoured cyanobacteria with altered gene inventories for coping with oxidative stress.     

Lineage-Specific Domain Fusion in the Evolution of Purine Nucleotide Cyclases in Cyanobacteria

Cyclic nucleotides (both cAMP and cGMP) play extremely important roles in cyanobacteria, such as regulating heterocyst formation, respiration, or gliding. Catalyzing the formation of cAMP and cGMP from ATP and GTP is a group of functionally important enzymes named adenylate cyclases and guanylate cyclases, respectively. To understand their evolutionary patterns, in this study, we presented a systematic analysis of all the cyclases in cyanobacterial genomes. We found that different cyanobacteria had various numbers of cyclases in view of their remarkable diversities in genome size and physiology. Most of these cyclases exhibited distinct domain architectures, which implies the versatile functions of cyanobacterial cyclases. Mapping the whole set of cyclase domain architectures from diverse prokaryotic organisms to their phylogenetic tree and detailed phylogenetic analysis of cyclase catalytic domains revealed that lineage-specific domain recruitment appeared to be the most prevailing pattern contributing to the great variability of cyanobacterial cyclase domain architectures. However, other scenarios, such as gene duplication, also occurred during the evolution of cyanobacterial cyclases. Sequence divergence seemed to contribute to the origin of putative guanylate cyclases which were found only in cyanobacteria. In conclusion, the comprehensive survey of cyclases in cyanobacteria provides novel insight into their potential evolutionary mechanisms and further functional implications.     

Cyanobacterial alkaline/neutral invertases. Origin of sucrose hydrolysis in the plant cytosol?

The aim of this work was to investigate the occurrence of invertase (Inv) in cyanobacteria. We describe the first isolation and characterization of prokaryotic alkaline/neutral Inv (A/N-Inv) genes. Two genes (invA and invB) were identified in Anabaena sp. PCC 7120, which share about 50-56% identity with plant A/N-Inv and encode proteins of about 53-55 kDa. The identification of these proteins was confirmed by biochemical and immunological studies with recombinant proteins and with the enzymes isolated from Anabaena cells. Expression analysis supported the important role of A/N-Inv in nitrogen-fixing growth conditions. Nevertheless, A/N-Inv activities were shown in all filamentous and unicellular cyanobacteria investigated, regardless of their capacity to fix dinitrogen. Searches in complete sequenced genomes showed that A/N-Inv homologues are restricted to cyanobacterial species and plants. In particular, filamentous nitrogen-fixing strains display two A/N-Inv genes and unicellular strains have only one. Phylogenetic analysis leads us to suggest that modern plant A/N-Inv might have originated from an orthologous ancestral gene after the endosymbiotic origin of chloroplasts.     

Cyanobacterial genomics for ecology and biotechnology

Cyanobacteria are the only prokaryotes that directly convert solar energy and CO(2) into organic matter by oxygenic photosynthesis, explaining their relevance for primary production in many ecosystems and the increasing interest for biotechnology. At present, there are more than 60 cyanobacteria for which a total genome sequence is publicly available. These cyanobacteria belong to different lifestyles and origins, coming from marine and freshwater aquatic environments, as well as terrestrial and symbiotic habitats. Genome sizes vary by a factor of six, from 1.44 Mb to 9.05 Mb, with the number of reported genes ranging from 1241 to 8462. Several studies have demonstrated how these sequences could be used to successfully infer important ecological, physiological and biotechnologically relevant characteristics. However, sequences of cyanobacterial origin also comprise a significant portion of certain metagenomes. Moreover, genome analysis has been employed for culture-independent approaches and for resequencing mutant strains, a very recent tool in cyanobacterial research.