RESUMO
Riboregulation has a major role in the fine-tuning of multiple bacterial processes. Among the RNA players, trans-encoded untranslated small RNAs (sRNAs) regulate complex metabolic networks by tuning expression from multiple target genes in response to numerous signals. In Sinorhizobium meliloti, over 400 sRNAs are expressed under different stimuli. The sRNA MmgR (standing for Makes more granules Regulator) has been of particular interest to us since its sequence and structure are highly conserved among the alphaproteobacteria and its expression is regulated by the amount and quality of the bacterium's available nitrogen source. In this work, we explored the biological role of MmgR in S. meliloti 2011 by characterizing the effect of a deletion of the internal conserved core of mmgR (mmgRΔ33-51). This mutation resulted in larger amounts of polyhydroxybutyrate (PHB) distributed into more intracellular granules than are found in the wild-type strain. This phenotype was expressed upon cessation of balanced growth owing to nitrogen depletion in the presence of surplus carbon (i.e., at a carbon/nitrogen molar ratio greater than 10). The normal PHB accumulation was complemented with a wild-type mmgR copy but not with unrelated sRNA genes. Furthermore, the expression of mmgR limited PHB accumulation in the wild type, regardless of the magnitude of the C surplus. Quantitative proteomic profiling and quantitative reverse transcription-PCR (qRT-PCR) revealed that the absence of MmgR results in a posttranscriptional overexpression of both PHB phasin proteins (PhaP1 and PhaP2). Together, our results indicate that the widely conserved alphaproteobacterial MmgR sRNA fine-tunes the regulation of PHB storage in S. melilotiIMPORTANCE High-throughput RNA sequencing has recently uncovered an overwhelming number of trans-encoded small RNAs (sRNAs) in diverse prokaryotes. In the nitrogen-fixing alphaproteobacterial symbiont of alfalfa root nodules Sinorhizobium meliloti, only four out of hundreds of identified sRNA genes have been functionally characterized. Thus, uncovering the biological role of sRNAs currently represents a major issue and one that is particularly challenging because of the usually subtle quantitative regulation contributed by most characterized sRNAs. Here, we have characterized the function of the broadly conserved alphaproteobacterial sRNA gene mmgR in S. meliloti Our results strongly suggest that mmgR encodes a negative regulator of the accumulation of polyhydroxybutyrate, the major carbon and reducing power storage polymer in S. meliloti cells growing under conditions of C/N overbalance.
Assuntos
Proteínas de Bactérias/metabolismo , Hidroxibutiratos/metabolismo , RNA Bacteriano/metabolismo , Sinorhizobium meliloti/metabolismo , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Carbono/metabolismo , Proteínas de Ligação a DNA/classificação , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , Mutação , Nitrogênio/metabolismo , Sinorhizobium meliloti/genéticaRESUMO
Streptomyces are Gram-positive microorganisms able to adapt and respond to different environmental conditions. It is the largest genus of Actinobacteria comprising over 900 species. During their lifetime, these microorganisms are able to differentiate, produce aerial mycelia and secondary metabolites. All of these processes are controlled by subtle and precise regulatory systems. Regulation at the transcriptional initiation level is probably the most common for metabolic adaptation in bacteria. In this mechanism, the major players are proteins named transcription factors (TFs), capable of binding DNA in order to repress or activate the transcription of specific genes. Some of the TFs exert their action just like activators or repressors, whereas others can function in both manners, depending on the target promoter. Generally, TFs achieve their effects by using one- or two-component systems, linking a specific type of environmental stimulus to a transcriptional response. After DNA sequencing, many streptomycetes have been found to have chromosomes ranging between 6 and 12Mb in size, with high GC content (around 70%). They encode for approximately 7000 to 10,000 genes, 50 to 100 pseudogenes and a large set (around 12% of the total chromosome) of regulatory genes, organized in networks, controlling gene expression in these bacteria. Among the sequenced streptomycetes reported up to now, the number of transcription factors ranges from 471 to 1101. Among these, 315 to 691 correspond to transcriptional regulators and 31 to 76 are sigma factors. The aim of this work is to give a state of the art overview on transcription factors in the genus Streptomyces.
Assuntos
Proteínas de Bactérias/fisiologia , Regulação Bacteriana da Expressão Gênica , Streptomyces/genética , Fatores de Transcrição/fisiologia , Proteínas de Bactérias/química , Proteínas de Bactérias/classificação , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/classificação , Proteínas de Ligação a DNA/fisiologia , Farmacorresistência Bacteriana/genética , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Redes e Vias Metabólicas/efeitos dos fármacos , Redes e Vias Metabólicas/genética , Células Procarióticas/fisiologia , Streptomyces/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/classificação , Xenobióticos/farmacologiaRESUMO
Characidae is the most species-rich family of freshwater fishes in the order Characiformes, with more than 1000 valid species that correspond to approximately 55% of the order. Few hypotheses about the composition and internal relationships within this family are available and most fail to reach an agreement. Among Characidae, Cheirodontinae is an emblematic group that includes 18 genera (1 fossil) and approximately 60 described species distributed throughout the Neotropical region. The taxonomic and systematic history of Cheirodontinae is complex, and only two hypotheses about the internal relationships in this subfamily have been reported to date. In the present study, we test the composition and relationships of fishes assigned to Cheirodontinae based on a broad taxonomic sample that also includes some characid incertae sedis taxa that were previously considered to be part of Cheirodontinae. We present phylogenetic analyses of a large molecular dataset of mitochondrial and nuclear DNA sequences. Our results reject the monophyly of Cheirodontinae as previously conceived, as well as the tribes Cheirodontini and Compsurini, and the genera Cheirodon, Compsura, Leptagoniates, Macropsobrycon, Odontostilbe, and Serrapinnus. On the basis of these results we propose: (1) the exclusion of Amazonspinther and Spintherobolus from the subfamily Cheirodontinae since they are the sister-group of all remaining Characidae; (2) the removal of Macropsobrycon xinguensis of the genus Macropsobrycon; (3) the removal of Leptagoniates pi of the genus Leptagoniates; (4) the inclusion of Leptagoniates pi in the subfamily Cheirodontinae; (5) the removal of Cheirodon stenodon of the genus Cheirodon and its inclusion in the subfamily Cheirodontinae under a new genus name; (6) the need to revise the polyphyletic genera Compsura, Odontostilbe, and Serrapinnus; and (7) the division of Cheirodontinae in three newly defined monophyletic tribes: Cheirodontini, Compsurini, and Pseudocheirodontini. Our results suggest that our knowledge about the largest Neotropical fish family, Characidae, still is incipient.
Assuntos
Núcleo Celular/genética , Characidae/classificação , DNA Mitocondrial/classificação , Mitocôndrias/genética , Filogenia , RNA Ribossômico 16S/classificação , Animais , Núcleo Celular/química , Characidae/genética , Citocromos b/classificação , Citocromos b/genética , DNA Mitocondrial/genética , Proteínas de Ligação a DNA/classificação , Proteínas de Ligação a DNA/genética , Água Doce , Especiação Genética , Proteínas de Homeodomínio/classificação , Proteínas de Homeodomínio/genética , Mitocôndrias/química , Cadeias Pesadas de Miosina/classificação , Cadeias Pesadas de Miosina/genética , RNA Ribossômico 16S/genéticaRESUMO
Acidithiobacillus ferrooxidans is a chemolithoautotrophic acidophilic bacterium that obtains its energy from the oxidation of ferrous iron, elemental sulfur, or reduced sulfur minerals. This capability makes it of great industrial importance due to its applications in biomining. During the industrial processes, A. ferrooxidans survives to stressing circumstances in its environment, such as an extremely acidic pH and high concentration of transition metals. In order to gain insight into the organization of A. ferrooxidans regulatory networks and to provide a framework for further studies in bacterial growth under extreme conditions, we applied a genome-wide annotation procedure to identify 87 A. ferrooxidans transcription factors. We classified them into 19 families that were conserved among diverse prokaryotic phyla. Our annotation procedure revealed that A. ferrooxidans genome contains several members of the ArsR and MerR families, which are involved in metal resistance and detoxification. Analysis of their sequences revealed known and potentially new mechanism to coordinate gene-expression in response to metal availability. A. ferrooxidans inhabit some of the most metal-rich environments known, thus transcription factors identified here seem to be good candidates for functional studies in order to determine their physiological roles and to place them into A. ferrooxidans transcriptional regulatory networks.
Assuntos
Acidithiobacillus/genética , Acidithiobacillus/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação Bacteriana da Expressão Gênica , Metais/metabolismo , Fatores de Transcrição/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Sítios de Ligação , Proteínas de Ligação a DNA/classificação , Proteínas de Ligação a DNA/genética , Homeostase , Dados de Sequência Molecular , Filogenia , Alinhamento de Sequência , Fatores de Transcrição/classificação , Fatores de Transcrição/genéticaRESUMO
Thermodimorphic fungi include most causative agents of systemic mycoses, but the molecular mechanisms that underlie their defining trait, i.e. the ability to shift between mould and yeast on temperature change alone, remain poorly understood. We hypothesised that the heat shock factor (Hsf), a protein that evolved to sense thermal stimuli quickly, might play a role in this process in addition to the known regulator Drk1 and the Ryp proteins. To test this hypothesis, we characterised the Hsf from the thermodimorph Paracoccidioides lutzii (formerly Paracoccidioides brasiliensis isolate 01). We show in the present work that PlHsf possesses regulatory domains that are exclusive of the Eurotiomycetidae family, suggesting evolutionary specialisation; that it can successfully rescue the otherwise lethal loss of the native protein of Saccharomyces cerevisiae; and that its DNA-binding domain is able to recognise regulatory elements from the promoters of both Drk1 and Ryp1. An in silico screening of all 1 kb sequences upstream of P. lutzii ORFs revealed that 7% of them possess a heat shock element. This is the first description of a heat shock factor in a thermodimorphic fungus.
Assuntos
Proteínas de Choque Térmico/genética , Paracoccidioides/genética , Paracoccidioidomicose/microbiologia , Sequência de Bases , Proteínas de Ligação a DNA/classificação , Proteínas de Ligação a DNA/genética , Evolução Molecular , Proteínas de Choque Térmico/classificação , Humanos , Dados de Sequência Molecular , Paracoccidioides/fisiologia , Filogenia , Regiões Promotoras Genéticas , Sequências Reguladoras de Ácido Nucleico , Proteínas de Saccharomyces cerevisiae/classificação , Proteínas de Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Temperatura , Fatores de Transcrição/classificação , Fatores de Transcrição/genéticaRESUMO
The Protein-DNA Interface database (PDIdb) is a repository containing relevant structural information of Protein-DNA complexes solved by X-ray crystallography and available at the Protein Data Bank. The database includes a simple functional classification of the protein-DNA complexes that consists of three hierarchical levels: Class, Type and Subtype. This classification has been defined and manually curated by humans based on the information gathered from several sources that include PDB, PubMed, CATH, SCOP and COPS. The current version of the database contains only structures with resolution of 2.5 A or higher, accounting for a total of 922 entries. The major aim of this database is to contribute to the understanding of the main rules that underlie the molecular recognition process between DNA and proteins. To this end, the database is focused on each specific atomic interface rather than on the separated binding partners. Therefore, each entry in this database consists of a single and independent protein-DNA interface.We hope that PDIdb will be useful to many researchers working in fields such as the prediction of transcription factor binding sites in DNA, the study of specificity determinants that mediate enzyme recognition events, engineering and design of new DNA binding proteins with distinct binding specificity and affinity, among others. Finally, due to its friendly and easy-to-use web interface, we hope that PDIdb will also serve educational and teaching purposes.