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1.
BMC Evol Biol ; 19(1): 224, 2019 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-31818253

RESUMO

BACKGROUND: Eukaryotes acquired the trait of oxygenic photosynthesis through endosymbiosis of the cyanobacterial progenitor of plastid organelles. Despite recent advances in the phylogenomics of Cyanobacteria, the phylogenetic root of plastids remains controversial. Although a single origin of plastids by endosymbiosis is broadly supported, recent phylogenomic studies are contradictory on whether plastids branch early or late within Cyanobacteria. One underlying cause may be poor fit of evolutionary models to complex phylogenomic data. RESULTS: Using Posterior Predictive Analysis, we show that recently applied evolutionary models poorly fit three phylogenomic datasets curated from cyanobacteria and plastid genomes because of heterogeneities in both substitution processes across sites and of compositions across lineages. To circumvent these sources of bias, we developed CYANO-MLP, a machine learning algorithm that consistently and accurately phylogenetically classifies ("phyloclassifies") cyanobacterial genomes to their clade of origin based on bioinformatically predicted function-informative features in tRNA gene complements. Classification of cyanobacterial genomes with CYANO-MLP is accurate and robust to deletion of clades, unbalanced sampling, and compositional heterogeneity in input tRNA data. CYANO-MLP consistently classifies plastid genomes into a late-branching cyanobacterial sub-clade containing single-cell, starch-producing, nitrogen-fixing ecotypes, consistent with metabolic and gene transfer data. CONCLUSIONS: Phylogenomic data of cyanobacteria and plastids exhibit both site-process heterogeneities and compositional heterogeneities across lineages. These aspects of the data require careful modeling to avoid bias in phylogenomic estimation. Furthermore, we show that amino acid recoding strategies may be insufficient to mitigate bias from compositional heterogeneities. However, the combination of our novel tRNA-specific strategy with machine learning in CYANO-MLP appears robust to these sources of bias with high accuracy in phyloclassification of cyanobacterial genomes. CYANO-MLP consistently classifies plastids as late-branching Cyanobacteria, consistent with independent evidence from signature-based approaches and some previous phylogenetic studies.


Assuntos
Cianobactérias/genética , Eucariotos/citologia , Eucariotos/genética , Plastídeos/genética , Evolução Biológica , Modelos Biológicos , Fotossíntese , Filogenia , RNA de Transferência , Simbiose
2.
Environ Pollut ; 255(Pt 2): 113327, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31600703

RESUMO

Fish-associated antibiotic resistance genes (ARGs) have attracted increasing attention due to their potential risks to human beings via the food chain. However, data are scarce regarding the antibiotic resistance in fish themselves. Herein, the antibiotic resistance genes (ARGs) were assessed in the gut of four major Chinese freshwater carp (i.e., silver carp, grass carp, bighead carp, and crucian carp) from food retail markets. Results show that the abundances of target ARGs (e.g., tetA, tetO, tetQ, tetW, sulI, sulII, and blaTEM-1) and class 1 integrase (intI1) were in the range 9.4 × 10-6 - 1.6 × 10-1 and 6.7 × 10-5 - 5.2 × 10-2 gene copies per 16S rRNA gene, respectively. The sulI, sulII, and tetQ strongly correlated with silver and mercury resistance genes (e.g., silE and merR). The microbial taxa of fish gut could be partly separated among retail markets based on the PCA analysis. About 15.0% of the OTUs in fish gut were shared and 74.5% of the shared OTUs were identified as Acidobacteria, Bacteroidetes, Chloroflexi, Cyanobacteria, Firmicutes, and Proteobacteria. These phyla may constitute the core microbiota in the guts of the four Chinese freshwater carp. The possible ARG hosts were revealed based on the network analysis, and the presence of pathogen-associated resistant genera in fish gut highlights the need to fully understand their potential human health risks.


Assuntos
Carpas/microbiologia , Farmacorresistência Bacteriana/genética , Microbioma Gastrointestinal/efeitos dos fármacos , Alimentos Marinhos/microbiologia , Acidobacteria/efeitos dos fármacos , Acidobacteria/genética , Acidobacteria/isolamento & purificação , Animais , Antibacterianos/farmacologia , Bacteroidetes/efeitos dos fármacos , Bacteroidetes/genética , Bacteroidetes/isolamento & purificação , Chloroflexi/efeitos dos fármacos , Chloroflexi/genética , Chloroflexi/isolamento & purificação , Cianobactérias/efeitos dos fármacos , Cianobactérias/genética , Cianobactérias/isolamento & purificação , Firmicutes/efeitos dos fármacos , Firmicutes/genética , Firmicutes/isolamento & purificação , Água Doce/microbiologia , Microbioma Gastrointestinal/genética , Genes Bacterianos/genética , Metais Pesados/toxicidade , Proteobactérias/efeitos dos fármacos , Proteobactérias/genética , Proteobactérias/isolamento & purificação , RNA Ribossômico 16S/genética
3.
Biochim Biophys Acta Bioenerg ; 1860(11): 148084, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31520614

RESUMO

Photosynthetic [2Fe-2S] plant-type ferredoxins have a central role in electron transfer between the photosynthetic chain and various metabolic pathways. Several genes are coding for [2Fe2S] ferredoxins in cyanobacteria, with four in the thermophilic cyanobacterium Thermosynechococcus elongatus. The structure and functional properties of the major ferredoxin Fd1 are well known but data on the other ferredoxins are scarce. We report the structural and functional properties of a novel minor type ferredoxin, Fd2 of T. elongatus, homologous to Fed4 from Synechocystis sp. PCC 6803. Remarkably, the midpoint potential of Fd2, Em = -440 mV, is lower than that of Fd1, Em = -372 mV. However, while Fd2 can efficiently react with photosystem I or nitrite reductase, time-resolved spectroscopy shows that Fd2 has a very low capacity to reduce ferredoxin-NADP+ oxidoreductase (FNR). These unique Fd2 properties are discussed in relation with its structure, solved at 1.38 Šresolution. The Fd2 structure significantly differs from other known ferredoxins structures in loop 2, N-terminal region, hydrogen bonding networks and surface charge distributions. UV-Vis, EPR, and Mid- and Far-IR data also show that the electronic properties of the [2Fe2S] cluster of Fd2 and its interaction with the protein differ from those of Fd1 both in the oxidized and reduced states. The structural analysis allows to propose that valine in the motif Cys53ValAsnCys56 of Fd2 and the specific orientation of Phe72, explain the electron transfer properties of Fd2. Strikingly, the nature of these residues correlates with different phylogenetic groups of cyanobacterial Fds. With its low redox potential and its discrimination against FNR, Fd2 exhibits a unique capacity to direct efficiently photosynthetic electrons to metabolic pathways not dependent on FNR.


Assuntos
Proteínas de Bactérias/metabolismo , Cianobactérias/metabolismo , Ferredoxinas/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Cianobactérias/genética , Ferredoxinas/química , Ferredoxinas/genética , Ligações de Hidrogênio , Cinética , Modelos Moleculares , Filogenia , Alinhamento de Sequência
4.
Sci Total Environ ; 687: 1369-1380, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31412470

RESUMO

In order to evaluate the recurrence of toxic cyanobacterial blooms and to determine the survival capabilities of the resistance cells through time, a sedimentary core spanning 6700 years was drilled in the eutrophic Lake Aydat. A multiproxy approach (density, magnetic susceptibility, XRF, pollen and non-pollen palynomorph analyses), was used initially to determine the sedimentation model and the land uses around the lake. Comparison with the akinete count revealed that Nostocales cyanobacteria have been present in Lake Aydat over a six thousand year period. This long-term cyanobacterial recurrence also highlights the past presence of both the anaC and mcyB genes, involved in anatoxin-a and microcystin biosynthesis, respectively, throughout the core. The first appearance of cyanobacteria seems to be linked to the natural damming of the river, while the large increase in akinete density around 1800 cal.yr BP can be correlated with the intensification of human activities (woodland clearance, crop planting, grazing, etc.) in the catchment area of the lake, and marks the beginning of a long period of eutrophication. This first investigation into the viability and germination potential of cyanobacteria over thousands of years reveals the ability of intact akinetes to undergo cell divisions even after 1800 years of sedimentation, which is 10 times longer than previously observed. This exceptional cellular resistance, coupled with the long-term eutrophic conditions of this lake, could partly explain the past and current recurrences of cyanobacterial proliferations.


Assuntos
Cianobactérias/genética , Monitoramento Ambiental , Poluição da Água/estatística & dados numéricos , Eutrofização , Sedimentos Geológicos/microbiologia , Lagos/microbiologia , Microcistinas , Recidiva , Estações do Ano , Tropanos
5.
Phytochemistry ; 166: 112059, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31280092

RESUMO

The cyanobacterial phylum is currently divided into five subsections (I-V), with the latter two containing no or false-branching (nostocalean) and true-branching (stigonematalean) cyanobacteria. Although morphological traits (such as cellular division and secondary branches) clearly separate both types of heterocytous cyanobacteria, molecular evidence indicates that stigonematalean cyanobacteria (Subsection V) do not form a monophyletic group but instead are interspersed and nested within the nostocalean cyanobacteria (Subsection IV). To further resolve the phylogeny of heterocytous cyanobacteria, we here analyzed the distribution of heterocyte glycolipids (HGs) in the true-branching cyanobacterium Stigonema ocellatum SAG 48.90 (type genus of Subsection V) and compared it with the HG inventory of other stigonematalean and nostocalean cyanobacteria. The most dominant HGs in S. ocellatum SAG 48.90 were 1-(O-hexose)-27-keto-3,25-octacosanediol (HG28 keto-diol) and 1-(O-hexose)-3,25,27-octacosanetriol (HG28 triol), which together constituted ca. 94% of all HGs. In addition, 1-(O-hexose)-3-keto-27-octacosanols (HG28 keto-ols), 1-(O-hexose)-3,27-octacosanediols (HG28 diols), 1-(O-hexose)-3-keto-27,29-triacontanediol (HG30 keto-diol) and 1-(O-hexose)-3,27,29-triacontanetriol (HG30 triol) occurred in minor abundances. Heterocyte glycolipids previously reported to be unique for stigonematalean cyanobacteria, i.e. 1-(O-hexose)-3,29,31-dotriacontanetriols (HG32 triols) and 1-(O-hexose)-3-keto-29,31-dotriacontanediols (HG32 keto-diols), were not detected in S. ocellatum SAG 48.90. Comparison of the HG distribution pattern with those of other heterocytous cyanobacteria indicated that S. ocellatum SAG 48.90 is most closely related to the nostocalean families Rivulariaceae and Scytonemataceae, which is complementary to reconstructed 16S rRNA gene sequence phylogenies. Our HG-based data thus provides evidence for the polyphyly of stigonematalean cyanobacteria, independent from molecular approaches, and points to the need for a critical re-evaluation of the current taxonomy of heterocytous cyanobacteria.


Assuntos
Cianobactérias/classificação , Cianobactérias/metabolismo , Glicolipídeos/metabolismo , Filogenia , Cianobactérias/genética , RNA Ribossômico 16S/genética
6.
Sci Total Environ ; 689: 789-796, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31280161

RESUMO

The sesquiterpene geosmin, mainly originating from cyanobacteria, is considered one of the problematic odor compounds responsible for unpleasant-tasting and -smelling water episodes in freshwater supplies. The biochemistry and genetics of geosmin synthesis in cyanobacteria is well-elucidated and the geosmin synthase gene (geo) has been cloned and characterized in recent years. However, understanding the diversity, origin, and evolution of geo has been hindered by the limited availability of geo sequences to date. On the basis of the cloned geo sequences from16 filamentous geosmin-producing cyanobacterial species, representing 11 genera in Nostocales and Oscillatoriales, the diversity and evolution of geo in cyanobacteria was systematically analyzed in this study. Homologous alignment revealed that geo is highly conserved among the examined cyanobacterial species, with DNA sequence identities >0.72. Phylogenetic reconstruction and codon bias analysis based on geo suggest that cyanobacterial geo form a monophyletic branch with a common origin and ancestor for cyanobacteria, actinomycetes, and myxobacteria. The global ratio of nonsynonymous/synonymous nucleotide substitutions (dN/dS) was 0.125, which is substantially <1 and indicates strong purifying selection in the evolution of cyanobacterial geo. To add to further interest, horizontal gene transfer of cyanobacterial geo in evolutionary history was confirmed by the discovery of an incongruent coevolutionary relationship between geo and housekeeping genes 16S rDNA and rpoC. The present study enhances the fundamental understanding of cyanobacterial geo in diversity and evolution, and sheds light on the development of molecular assays for detection and molecular ecology research of geosmin-producing cyanobacteria.


Assuntos
Proteínas de Bactérias/genética , Cianobactérias/genética , Evolução Molecular , Variação Genética , Naftóis/metabolismo , Proteínas de Bactérias/metabolismo , Cianobactérias/metabolismo , DNA Bacteriano/análise , Filogenia , Análise de Sequência de DNA , Especificidade da Espécie
7.
Int J Mol Sci ; 20(11)2019 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-31181593

RESUMO

The slow but temperature-insensitive adenosine triphosphate (ATP) hydrolysis reaction in KaiC is considered as one of the factors determining the temperature-compensated period length of the cyanobacterial circadian clock system. Structural units responsible for this low but temperature-compensated ATPase have remained unclear. Although whole-KaiC scanning mutagenesis can be a promising experimental strategy, producing KaiC mutants and assaying those ATPase activities consume considerable time and effort. To overcome these bottlenecks for in vitro screening, we optimized protocols for expressing and purifying the KaiC mutants and then designed a high-performance liquid chromatography system equipped with a multi-channel high-precision temperature controller to assay the ATPase activity of multiple KaiC mutants simultaneously at different temperatures. Through the present protocol, the time required for one KaiC mutant is reduced by approximately 80% (six-fold throughput) relative to the conventional protocol with reasonable reproducibility. For validation purposes, we picked up three representatives from 86 alanine-scanning KaiC mutants preliminarily investigated thus far and characterized those clock functions in detail.


Assuntos
Proteínas de Bactérias/genética , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Cianobactérias/genética , Mutação , Adenosina Trifosfatases/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/química , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/metabolismo , Cianobactérias/metabolismo , Técnicas Genéticas
8.
Int J Mol Sci ; 20(12)2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-31208089

RESUMO

Cyanobacteria have cyanobacteriochromes (CBCRs), which are photoreceptors that bind to a linear tetrapyrrole chromophore and sense UV-to-visible light. A recent study revealed that the dual-Cys CBCR AM1_1186g2 covalently attaches to phycocyanobilin and exhibits unique photoconversion between a Pr form (red-absorbing dark state, λmax = 641 nm) and Pb form (blue-absorbing photoproduct, λmax = 416 nm). This wavelength separation is larger than those of the other CBCRs, which is advantageous for optical tools. Nowadays, bioimaging and optogenetics technologies are powerful tools for biological research. In particular, the utilization of far-red and near-infrared light sources is required for noninvasive applications to mammals because of their high potential to penetrate into deep tissues. Biliverdin (BV) is an intrinsic chromophore and absorbs the longest wavelength among natural linear tetrapyrrole chromophores. Although the BV-binding photoreceptors are promising platforms for developing optical tools, AM1_1186g2 cannot efficiently attach BV. Herein, by rationally introducing several replacements, we developed a BV-binding AM1_1186g2 variant, KCAP_QV, that exhibited reversible photoconversion between a Pfr form (far-red-absorbing dark state, λmax = 691 nm) and Pb form (λmax = 398 nm). This wavelength separation reached 293 nm, which is the largest among the known phytochrome and CBCR photoreceptors. In conclusion, the KCAP_QV molecule developed in this study can offer an alternative platform for the development of unique optical tools.


Assuntos
Biliverdina/química , Luz , Fitocromo/química , Fitocromo/metabolismo , Engenharia de Proteínas , Sequência de Aminoácidos , Biliverdina/metabolismo , Cianobactérias/genética , Cianobactérias/metabolismo , Estrutura Molecular , Fotorreceptores Microbianos/genética , Fotorreceptores Microbianos/metabolismo , Fitocromo/genética , Estabilidade Proteica
9.
Appl Biochem Biotechnol ; 189(4): 1127-1140, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31168708

RESUMO

Cyanobacteria have immense prospective as a platform for renewable energy; however, a major barrier in achieving optimal productivity is the low lipid yield. Fremyella diplosiphon, a model cyanobacterium, is an ideal biofuel agent due to its desirable fatty acid methyl esters (FAMEs). To enhance lipid content, we overexpressed the sterol desaturase (SD) gene in F. diplosiphon B481 wild type by genetic transformation. This effort resulted in a transformant (B481-SD) with a 64-fold increase in the SD gene at the mRNA transcript level, with no loss in growth and pigmentation. The transformant was persistently grown for over 32 generations indicating long-term stability and vitality. We observed 27.3% and 23% increases in total lipid content and unsaturated FAMEs respectively in B481-SD transesterified lipids with methyl octadecadienoate as the most abundant unsaturated component. In addition, we detected an 81% increase in FAME composition in the transformant compared with the wild type. Theoretical physical and chemical properties confirmed a FAME profile with very high cetane number (65.972-67.494) and oxidative stability (50.493-18.66 h) in the engineered strain. Results of the study offer a promising approach to augment F. diplosiphon total lipid content and unsaturated FAMEs, thus paving the way to enhance biofuel capacity of the organism.


Assuntos
Biocombustíveis , Cianobactérias , Ácidos Graxos Insaturados , Engenharia Metabólica , Cianobactérias/genética , Cianobactérias/metabolismo , Ésteres/metabolismo , Ácidos Graxos Insaturados/biossíntese , Ácidos Graxos Insaturados/genética
10.
Biochim Biophys Acta Bioenerg ; 1860(7): 549-561, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31173730

RESUMO

Phycoerythrin (PE) present in the distal ends of light-harvesting phycobilisome rods in Fremyella diplosiphon (Tolypothrix sp. PCC 7601) contains five phycoerythrobilin (PEB) chromophores attached to six cysteine residues for efficient green light capture for photosynthesis. Chromophore ligation on PE subunits occurs through bilin lyase catalyzed reactions, but the characterization of the roles of all bilin lyases for phycoerythrin is not yet complete. To gain a more complete understanding about the individual functions of CpeZ and CpeY in PE biogenesis in cyanobacteria, we examined PE and phycobilisomes purified from wild type F. diplosiphon, cpeZ and cpeY knockout mutants. We find that the cpeZ and cpeY mutants accumulate less PE than wild type cells. We show that in the cpeZ mutant, chromophorylation of both PE subunits is affected, especially the Cys-80 and Cys-48/Cys-59 sites of CpeB, the beta-subunit of PE. The cpeY mutant showed reduced chromophorylation at Cys-82 of CpeA. We also show that, in vitro, CpeZ stabilizes PE subunits and assists in refolding of CpeB after denaturation. Taken together, we conclude that CpeZ acts as a chaperone-like protein, assisting in the folding/stability of PE subunits, allowing bilin lyases such as CpeY and CpeS to attach PEB to their PE subunit.


Assuntos
Proteínas de Bactérias/metabolismo , Cianobactérias/metabolismo , Chaperonas Moleculares/metabolismo , Ficoeritrina/biossíntese , Proteínas Recombinantes/metabolismo , Proteínas de Bactérias/genética , Cianobactérias/genética , Cianobactérias/crescimento & desenvolvimento , Mutação , Proteínas Recombinantes/genética
11.
Genome Biol Evol ; 11(7): 1959-1964, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31243438

RESUMO

Plant endosymbiosis with nitrogen-fixing cyanobacteria has independently evolved in diverse plant lineages, offering a unique window to study the evolution and genetics of plant-microbe interaction. However, very few complete genomes exist for plant cyanobionts, and therefore little is known about their genomic and functional diversity. Here, we present four complete genomes of cyanobacteria isolated from bryophytes. Nanopore long-read sequencing allowed us to obtain circular contigs for all the main chromosomes and most of the plasmids. We found that despite having a low 16S rRNA sequence divergence, the four isolates exhibit considerable genome reorganizations and variation in gene content. Furthermore, three of the four isolates possess genes encoding vanadium (V)-nitrogenase (vnf), which is uncommon among diazotrophs and has not been previously reported in plant cyanobionts. In two cases, the vnf genes were found on plasmids, implying possible plasmid-mediated horizontal gene transfers. Comparative genomic analysis of vnf-containing cyanobacteria further identified a conserved gene cluster. Many genes in this cluster have not been functionally characterized and would be promising candidates for future studies to elucidate V-nitrogenase function and regulation.


Assuntos
Cianobactérias/genética , Família Multigênica/genética , Nitrogenase/genética , Nitrogenase/classificação , Filogenia , Plasmídeos/genética , RNA Ribossômico 16S/genética
12.
Mar Pollut Bull ; 142: 135-144, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31232286

RESUMO

This study investigated the microbial structure in the surface seawater from five coastal sites around Xiamen Island, China, over four seasons to evaluate seasonal environmental fluctuations impact on them. This subtropical island is characterized by long, hot, humid summers, and short, mild, dry winters. All sites were dominated by Proteobacteria, Bacteroidetes, Cyanobacteria, and Firmicutes; microbial community composition was similar across four seasons. However, larger proportions of Gammaproteobacteria and Bacillus were observed during the summer than during any other season. The high ratio of Bacillus, Bacteroidetes, and Clostridia richness to Alphaproteobacteria richness in the summer, suggested that the sites we tested were heavily affected by waste water to other seasons. Correlation-based network analyses among the bacterial species and environmental variables indicated important connections between physiochemical variables and specific taxonomic groups. Collectively, our results suggested that seasonal shifts and wastewater pollution together shape the structures of the microbial communities around Xiamen Island.


Assuntos
Bactérias/genética , Água do Mar/microbiologia , China , Cianobactérias/genética , Estuários , Gammaproteobacteria/genética , Ilhas , Filogenia , Proteobactérias/genética , RNA Ribossômico 16S/genética , Estações do Ano , Águas Residuárias , Microbiologia da Água , Poluição da Água
13.
Sheng Wu Gong Cheng Xue Bao ; 35(5): 795-804, 2019 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-31222998

RESUMO

The cyanobacterial circadian clock has three relatively independent parts: the input path, the core oscillator, and the output path. The core oscillator is composed of three clock proteins: KaiA, KaiB, and KaiC. The interactions among these three proteins generate a rhythmic signal and convey the input signals to the output signals to maintain the accuracy and stability of the oscillation of downstream signals. Based on the cyanobacterial circadian clock and the structure, function, and interaction of the clock proteins of the core oscillator, combining the recent results from our laboratory, this review summarized the recent progresses of the molecular mechanism of KaiA in regulating KaiC's enzymatic activity, mediating phase reset of the oscillator, and competing with CikA for the binding site of KaiB.


Assuntos
Proteínas de Bactérias , Relógios Circadianos , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano , Cianobactérias , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Relógios Circadianos/genética , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/metabolismo , Cianobactérias/genética , Ativação Enzimática/genética
14.
Mar Drugs ; 17(5)2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31067786

RESUMO

Microcystins are a family of chemically diverse hepatotoxins produced by distantly related cyanobacteria and are potent inhibitors of eukaryotic protein phosphatases 1 and 2A. Here we provide evidence for the biosynthesis of rare variants of microcystin that contain a selection of homo-amino acids by the benthic strain Phormidium sp. LP904c. This strain produces at least 16 microcystin chemical variants many of which contain homophenylalanine or homotyrosine. We retrieved the complete 54.2 kb microcystin (mcy) gene cluster from a draft genome assembly. Analysis of the substrate specificity of McyB1 and McyC adenylation domain binding pockets revealed divergent substrate specificity sequences, which could explain the activation of homo-amino acids which were present in 31% of the microcystins detected and included variants such as MC-LHty, MC-HphHty, MC-LHph and MC-HphHph. The mcy gene cluster did not encode enzymes for the synthesis of homo-amino acids but may instead activate homo-amino acids produced during the synthesis of anabaenopeptins. We observed the loss of microcystin during cultivation of a closely related strain, Phormidium sp. DVL1003c. This study increases the knowledge of benthic cyanobacterial strains that produce microcystin variants and broadens the structural diversity of known microcystins.


Assuntos
Cianobactérias/genética , Cianobactérias/metabolismo , Microcistinas/biossíntese , Microcistinas/genética , Sequência de Aminoácidos , Aminoácidos/metabolismo , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Genes Bacterianos , Microcistinas/química , Família Multigênica , Filogenia , Domínios e Motivos de Interação entre Proteínas , Análise de Sequência de DNA
15.
Int J Mol Sci ; 20(9)2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-31071918

RESUMO

BACKGROUND: Peanut (Arachis hypogaea L.), an important oilseed and food legume, is widely cultivated in the semi-arid tropics. Drought is the major stress in this region which limits productivity. Microbial communities in the rhizosphere are of special importance to stress tolerance. However, relatively little is known about the relationship between drought and microbial communities in peanuts. METHOD: In this study, deep sequencing of the V3-V4 region of the 16S rRNA gene was performed to characterize the microbial community structure of drought-treated and untreated peanuts. RESULTS: Taxonomic analysis showed that Actinobacteria, Proteobacteria, Saccharibacteria, Chloroflexi, Acidobacteria and Cyanobacteria were the dominant phyla in the peanut rhizosphere. Comparisons of microbial community structure of peanuts revealed that the relative abundance of Actinobacteria and Acidobacteria dramatically increased in the seedling and podding stages in drought-treated soil, while that of Cyanobacteria and Gemmatimonadetes increased in the flowering stage in drought-treated rhizospheres. Metagenomic profiling indicated that sequences related to metabolism, signaling transduction, defense mechanism and basic vital activity were enriched in the drought-treated rhizosphere, which may have implications for plant survival and drought tolerance. CONCLUSION: This microbial communities study will form the foundation for future improvement of drought tolerance of peanuts via modification of the soil microbes.


Assuntos
Arachis/microbiologia , Secas , Microbiota/genética , Estresse Fisiológico/genética , Acidobacteria/classificação , Acidobacteria/genética , Actinobacteria/classificação , Actinobacteria/genética , Arachis/genética , Chloroflexi/classificação , Chloroflexi/genética , Cianobactérias/classificação , Cianobactérias/genética , Sequenciamento de Nucleotídeos em Larga Escala , Filogenia , Raízes de Plantas/microbiologia , Proteobactérias/classificação , Proteobactérias/genética , RNA Ribossômico 16S/genética , Rizosfera , Plântula/genética , Microbiologia do Solo , Clima Tropical
16.
mBio ; 10(3)2019 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088928

RESUMO

Marine sponges are recognized as valuable sources of bioactive metabolites and renowned as petri dishes of the sea, providing specialized niches for many symbiotic microorganisms. Sponges of the family Dysideidae are well documented to be chemically talented, often containing high levels of polyhalogenated compounds, terpenoids, peptides, and other classes of bioactive small molecules. This group of tropical sponges hosts a high abundance of an uncultured filamentous cyanobacterium, Hormoscilla spongeliae Here, we report the comparative genomic analyses of two phylogenetically distinct Hormoscilla populations, which reveal shared deficiencies in essential pathways, hinting at possible reasons for their uncultivable status, as well as differing biosynthetic machinery for the production of specialized metabolites. One symbiont population contains clustered genes for expanded polybrominated diphenylether (PBDE) biosynthesis, while the other instead harbors a unique gene cluster for the biosynthesis of the dysinosin nonribosomal peptides. The hybrid sequencing and assembly approach utilized here allows, for the first time, a comprehensive look into the genomes of these elusive sponge symbionts.IMPORTANCE Natural products provide the inspiration for most clinical drugs. With the rise in antibiotic resistance, it is imperative to discover new sources of chemical diversity. Bacteria living in symbiosis with marine invertebrates have emerged as an untapped source of natural chemistry. While symbiotic bacteria are often recalcitrant to growth in the lab, advances in metagenomic sequencing and assembly now make it possible to access their genetic blueprint. A cell enrichment procedure, combined with a hybrid sequencing and assembly approach, enabled detailed genomic analysis of uncultivated cyanobacterial symbiont populations in two chemically rich tropical marine sponges. These population genomes reveal a wealth of secondary metabolism potential as well as possible reasons for historical difficulties in their cultivation.


Assuntos
Cianobactérias/genética , Metagenômica , Poríferos/microbiologia , Simbiose/genética , Animais , Produtos Biológicos/metabolismo , Cianobactérias/metabolismo , Genômica , Éteres Difenil Halogenados/metabolismo , Indóis/metabolismo , Família Multigênica , Filogenia , Pirróis/metabolismo , Clima Tropical
17.
mBio ; 10(3)2019 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-31113897

RESUMO

The biosynthesis of the unique cyanobacterial (oxyphotobacterial) indole-phenolic UVA sunscreen, scytonemin, is coded for in a conserved operon that contains both core metabolic genes and accessory, aromatic amino acid biosynthesis genes dedicated to supplying scytonemin's precursors. Comparative genomics shows conservation of this operon in many, but not all, cyanobacterial lineages. Phylogenetic analyses of the operon's aromatic amino acid genes indicate that five of them were recruited into the operon after duplication events of their respective housekeeping cyanobacterial cognates. We combined the fossil record of cyanobacteria and relaxed molecular clock models to obtain multiple estimates of these duplication events, setting a minimum age for the evolutionary advent of scytonemin at 2.1 ± 0.3 billion years. The same analyses were used to estimate the advent of cyanobacteria as a group (and thus the appearance of oxygenic photosynthesis), at 3.6 ± 0.2 billion years before present. Post hoc interpretation of 16S rRNA-based Bayesian analyses was consistent with these estimates. Because of physiological constraints on the use of UVA sunscreens in general, and the biochemical constraints of scytonemin in particular, scytonemin's age must postdate the time when Earth's atmosphere turned oxic, known as the Great Oxidation Event (GOE). Indeed, our biological estimate is in agreement with independent geochemical estimates for the GOE. The difference between the estimated ages of oxygenic photosynthesis and the GOE indicates the long span (on the order of a billion years) of the era of "oxygen oases," when oxygen was available locally but not globally.IMPORTANCE The advent of cyanobacteria, with their invention of oxygenic photosynthesis, and the Great Oxidation Event are arguably among the most important events in the evolutionary history of life on Earth. Oxygen is a significant toxicant to all life, but its accumulation in the atmosphere also enabled the successful development and proliferation of many aerobic organisms, especially metazoans. The currently favored dating of the Great Oxidation Event is based on the geochemical rock record. Similarly, the advent of cyanobacteria is also often drawn from the same estimates because in older rocks paleontological evidence is scarce or has been discredited. Efforts to obtain molecular evolutionary alternatives have offered widely divergent estimates. Our analyses provide a novel means to circumvent these limitations and allow us to estimate the large time gap between the two events.


Assuntos
Vias Biossintéticas/genética , Cianobactérias/genética , Cianobactérias/metabolismo , Evolução Molecular , Indóis/metabolismo , Fenóis/metabolismo , Filogenia , Protetores Solares/metabolismo , Fósseis
18.
Planta Med ; 85(11-12): 973-980, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31026875

RESUMO

Abietane diterpenoids, containing a quinone moiety, are synthesized in the roots of several Salvia species. Promising cytotoxicity and antiproliferative activities have been reported for these compounds in various cell and animal models. We have recently shown that aethiopinone, an o-naphto-quinone diterpene, produced in the roots of different Salvia species, is selectively cytotoxic against the A375 melanoma cell line. To enhance the synthesis of this abietane diterpenoid, we have engineered the plastidial 2-C-methyl-D-erythritol 4-phosphate-derived isoprenoid pathway in Salvia sclarea hairy roots by ectopic expression and plastid targeting of cyanobacterial genes encoding the 1-deoxy-D-xylulose 5-phosphate synthase or 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene, the first two enzymatic steps of the plastidial MEP pathway, from which plant diterpenes primarily derive. Plastid-targeted expression of 1-deoxy-D-xylulose 5-phosphate synthase and 1-deoxy-D-xylulose-5-phosphate reductoisomerase proteins significantly enhanced the yield of aethiopinone by a 3-fold and about 6-fold increase, respectively. The accumulation of other abietane-type diterpenes (ferruginol, salvipisone, and carnosic acid), with interesting antiproliferative activity, was also increased. Compared to our previous data obtained by overexpressing the plant orthologous 1-deoxy-D-xylulose 5-phosphate synthase and 1-deoxy-D-xylulose-5-phosphate reductoisomerase genes in S. sclarea hairy roots, the results presented here confirm that the bacterial 1-deoxy-D-xylulose-5-phosphate reductoisomerase enzyme plays a major role than the DXS enzyme in the biosynthetic pathway of this class of compounds and that its ectopic expression does not conflict with active hairy root growth, resulting in a balanced trade-off between the transgenic hairy root final biomass and the increased content of o-naphto-quinone diterpenes, with interesting biological activities.


Assuntos
/metabolismo , Cianobactérias/genética , Genes Bacterianos/genética , Raízes de Plantas/metabolismo , Salvia/metabolismo , /análise , Western Blotting , Regulação da Expressão Gênica de Plantas , Raízes de Plantas/química , Raízes de Plantas/genética , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase , Salvia/química , Salvia/genética
19.
J Microbiol ; 57(6): 450-460, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31012060

RESUMO

Next-generation DNA sequencing technology was applied to generate molecular data from semiarid reservoirs during well-defined seasons. Target sequences of 16S-23S rRNA ITS and cpcBA-IGS were used to reveal the taxonomic groups of cyanobacteria present in the samples, and genes coding for cyanotoxins such as microcystins (mcyE), saxitoxins (sxtA), and cylindrospermopsins (cyrJ) were investigated. The presence of saxitoxins in the environmental samples was evaluated using ELISA kit. Taxonomic analyses of high-throughput DNA sequencing data showed the dominance of the genus Microcystis in Mundaú reservoir. Furthermore, it was the most abundant genus in the dry season in Ingazeira reservoir. In the rainy season, 16S-23S rRNA ITS analysis revealed that Cylindrospermopsis raciborskii comprised 46.8% of the cyanobacterial community in Ingazeira reservoir, while the cpcBAIGS region revealed that C. raciborskii (31.8%) was the most abundant taxon followed by Sphaerospermopsis aphanizomenoides (17.3%) and Planktothrix zahidii (16.6%). Despite the presence of other potential toxin-producing genera, the detected sxtA gene belonged to C. raciborskii, while the mcyE gene belonged to Microcystis in both reservoirs. The detected mcyE gene had good correlation with MC content, while the amplification of the sxtA gene was related to the presence of STX. The cyrJ gene was not detected in these samples. Using DNA analyses, our results showed that the cyanobacterial composition of Mundaú reservoir was similar in successive dry seasons, and it varied between seasons in Ingazeira reservoir. In addition, our data suggest that some biases of analysis influenced the cyanobacterial communities seen in the NGS output of Ingazeira reservoir.


Assuntos
Biodiversidade , Cianobactérias/classificação , Cianobactérias/isolamento & purificação , Água Potável/microbiologia , Análise de Sequência de DNA/métodos , Microbiologia da Água , Abastecimento de Água , Toxinas Bacterianas/análise , Toxinas Bacterianas/genética , Toxinas Bacterianas/isolamento & purificação , Brasil , Cianobactérias/genética , DNA Bacteriano/análise , Monitoramento Ambiental/métodos , Genes Bacterianos/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Metagenômica/métodos , Microcystis/genética , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Saxitoxina/genética , Estações do Ano , Uracila/análogos & derivados
20.
Environ Sci Pollut Res Int ; 26(16): 16097-16104, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30968298

RESUMO

The phyllosphere provides appropriate conditions for colonization by microorganisms, including diazotrophic bacteria. However, a poor understanding of the effects of the atmospheric environment on the phyllospheric diazotrophic communities persists. We detected the biodiversity, abundance, and activity of nitrogen-fixing bacteria in the phyllospheres of two evergreen shrubs, Nerium indicum Mill. and Osmanthus sp., sampled from urban areas with heavy traffic, a college campus, and a forest. Quantitative PCR analysis indicated that the copy numbers of nifH sequences were highest in the phyllospheres of both plants in heavy-traffic urban areas and correlated with the recorded nitrogenase activities of the phyllospheres. Similarly, the phyllosphere from heavy-traffic urban areas also possessed the highest biodiversity indices of diazotrophic communities from both the two plants. Pyrosequencing analysis revealed a diversity of nifH sequences in phyllosphere that were mostly uniquely found in the phyllosphere, and many of these were proteobacteria-like and cyanobacteria-like. Members of the Proteobacteria, mostly of which were not closely related to unknown organisms, were detected exclusively in the phyllosphere and represented substantial fractions of their associated diazotrophic communities. Our study provides initial insight into the shifts in the biodiversity and community structure of N2-fixing microorganisms in the phyllospheres of different atmospheric environments.


Assuntos
Nerium/microbiologia , Bactérias Fixadoras de Nitrogênio/fisiologia , Oleaceae/microbiologia , Biodiversidade , Cianobactérias/genética , Ecossistema , Florestas , Oxirredutases/genética , Proteobactérias/genética , Reação em Cadeia da Polimerase em Tempo Real , Urbanização
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