Enterotype-Dependent Probiotic-Mediated Changes in the Male Rat Intestinal Microbiome In Vivo and In Vitro.

Beneficial properties of lactic acid bacteria have been known long ago, but particular interest in probiotics has arisen in the last two decades due to the understanding of the important role of intestinal microflora in human life. Thus, the ability of probiotics to support healthy homeostasis of gut microbiomes has received particular attention. Here, we evaluated the effect of a probiotic consisting of and on the gut microbiome of male rats, assessed their persistence in the fecal biota, and compared probiotic-mediated changes in vitro and in vivo.

Neuromodulators as Interdomain Signaling Molecules Capable of Occupying Effector Binding Sites in Bacterial Transcription Factors.

Hormones and neurotransmitters are important components of inter-kingdom signaling systems that ensure the coexistence of eukaryotes with their microbial community. Their ability to affect bacterial physiology, metabolism, and gene expression was evidenced by various experimental approaches, but direct penetration into bacteria has only recently been reported. This opened the possibility of considering neuromodulators as potential effectors of bacterial ligand-dependent regulatory proteins.

The Fate and Functionality of Alien tRNA Fragments in Culturing Medium and Cells of .

Numerous observations have supported the idea that various types of noncoding RNAs, including tRNA fragments (tRFs), are involved in communications between the host and its microbial community. The possibility of using their signaling function has stimulated the study of secreted RNAs, potentially involved in the interspecies interaction of bacteria. This work aimed at identifying such RNAs and characterizing their maturation during transport.

Sense and antisense RNA products of the gene can affect motility and chemotaxis acting independent of the UxuR protein.

Small non-coding and antisense RNAs are widespread in all kingdoms of life, however, the diversity of their functions in bacteria is largely unknown. Here, we study RNAs synthesised from divergent promoters located in the 3'-end of the gene, encoding transcription factor regulating hexuronate metabolism in These overlapping promoters were predicted with rather high scores, effectively bound RNA polymerase and and were capable of initiating transcription in sense and antisense directions. The genome-wide correlation between promoter scores and RNA polymerase binding and was higher for promoters located on the antisense strands of the genes, however, sense promoters within the gene were more active.

Differential Impact of Hexuronate Regulators ExuR and UxuR on the Proteome.

ExuR and UxuR are paralogous proteins belonging to the GntR family of transcriptional regulators. Both are known to control hexuronic acid metabolism in a variety of Gammaproteobacteria but the relative impact of each of them is still unclear. Here, we apply 2D difference electrophoresis followed by mass-spectrometry to characterise the changes in the proteome in response to a or deletion.

: Occurrence in the Human Gut Microbiota and -Mer-Based Assessment of Intraspecies Diversity.

Alignment-free approaches employing short -mers as barcodes for individual genomes have created a new strategy for taxonomic analysis and paved a way for high-resolution phylogeny. Here, we introduce this strategy for the species as a taxon requiring barcoding support for precise systematics. Using this approach for phylotyping of VKM B-1144 at the genus level, we identified four phylogroups and found that 12A belongs to one of them, rather than to the clade.

Suppression of Growth Dynamics via RNAs Secreted by Competing Bacteria.

With the discovery of secreted RNAs, it has become apparent that the biological role of regulatory oligonucleotides likely goes beyond the borders of individual cells. However, the mechanisms of their action are still comprehended only in general terms and mainly for eukaryotic microRNAs, which can interfere with mRNAs even in distant recipient cells. It has recently become clear that bacterial cells lacking interference systems can also respond to eukaryotic microRNAs that have targets in their genomes.

Excessive Promoters as Silencers of Genes Horizontally Acquired by .

Horizontally acquired genes are usually transcriptionally inactive, although most of them are associated with genomic loci enriched with promoter-like sequences forming "promoter islands." We hypothesized that lateral DNA transfer induces local mutagenesis, accumulating AT base pairs and creating promoter-like sequences, whose occupancy with RNA polymerase and a specific silencer H-NS suppresses the transcription of foreign genes. Error-prone mutagenesis was implemented for the "promoter island" of a foreign gene and the promoter region of an inherent gene .

Unique -mers as Strain-Specific Barcodes for Phylogenetic Analysis and Natural Microbiome Profiling.

The need for a comparative analysis of natural metagenomes stimulated the development of new methods for their taxonomic profiling. Alignment-free approaches based on the search for marker -mers turned out to be capable of identifying not only species, but also strains of microorganisms with known genomes. Here, we evaluated the ability of genus-specific -mers to distinguish eight phylogroups of (A, B1, C, E, D, F, G, B2) and assessed the presence of their unique 22-mers in clinical samples from microbiomes of four healthy people and four patients with Crohn's disease.

Between computational predictions and high-throughput transcriptional profiling: in depth expression analysis of the OppB trans-membrane subunit of Escherichia coli OppABCDF oligopeptide transporter.

Bacterial oligopeptide transporters encoded by arrays of opp genes are implicated in a wide variety of physiological functions including nutrient acquisition, cell-to-cell communication, host-pathogen interaction. Combining the five opp genes in one oppABCDF operon of Escherichia coli assumes unified principle of their transcriptional regulation, which should provide a comparable amounts of translated products. This, however, contradicts the experimentally detected disproportion in the abundance of periplasmic OppA and the trans-membrane subunits OppB and OppC.

Overproduction and purification of the Escherichia coli transcription factors "toxic" to a bacterial cell.

Transcription factors play a crucial role in control of life of a bacterial cell, working as switchers to a different life style or pathogenicity. To reconstruct the network of regulatory events taking place in changing growth conditions, we need to know regulons of as many transcription factors as possible, and motifs recognized by them. Experimentally this can be attained via ChIP-seq in vivo, SELEX and DNAse I footprinting in vitro.

A cohabiting bacterium alters the spectrum of short RNAs secreted by Escherichia coli.

Recently, it has been found that bacteria secrete short RNAs able to affect gene expression in eukaryotic cells, while certain mammalian microRNAs shape the gut microbiome altering bacterial transcriptome. The involvement of bacterial RNAs in communication with other bacteria is also expected, but has not been documented yet. Here, we compared the fractions of extremely short (12-22 nucleotides) RNAs secreted by Escherichia coli grown in a pure culture and jointly with bacteria of the Paenibacillus genus.

The Oligomeric Form of the Escherichia coli Dps Protein Depends on the Availability of Iron Ions.

The Dps protein of , which combines ferroxidase activity and the ability to bind DNA, is effectively used by bacteria to protect their genomes from damage. Both activities depend on the integrity of this multi-subunit protein, which has an inner cavity for iron oxides; however, the diversity of its oligomeric forms has only been studied fragmentarily. Here, we show that iron ions stabilize the dodecameric form of Dps.

The nucleoid protein Dps binds genomic DNA of Escherichia coli in a non-random manner.

Dps is a multifunctional homododecameric protein that oxidizes Fe2+ ions accumulating them in the form of Fe2O3 within its protein cavity, interacts with DNA tightly condensing bacterial nucleoid upon starvation and performs some other functions. During the last two decades from discovery of this protein, its ferroxidase activity became rather well studied, but the mechanism of Dps interaction with DNA still remains enigmatic. The crucial role of lysine residues in the unstructured N-terminal tails led to the conventional point of view that Dps binds DNA without sequence or structural specificity.

Control of hexuronate metabolism in Escherichia coli by the two interdependent regulators, ExuR and UxuR: derepression by heterodimer formation.

Two homologous proteins, UxuR and ExuR, were previously predicted to repress synthesis of enzymes required for hexuronic acid metabolism, but little is known about the relative roles of these proteins in gene regulation. We confirmed the previous report that UxuR is essential for rapid growth with d-glucuronate as the primary source of carbon and energy. In contrast, an exuR mutant grew more rapidly on d-glucuronate than the parent.

Translatomics combined with transcriptomics and proteomics reveals novel functional, recently evolved orphan genes in Escherichia coli O157:H7 (EHEC).

Background: Genomes of E. coli, including that of the human pathogen Escherichia coli O157:H7 (EHEC) EDL933, still harbor undetected protein-coding genes which, apparently, have escaped annotation due to their small size and non-essential function. To find such genes, global gene expression of EHEC EDL933 was examined, using strand-specific RNAseq (transcriptome), ribosomal footprinting (translatome) and mass spectrometry (proteome).

Structural modeling of the ExuR and UxuR transcription factors of E. coli: search for the ligands affecting their regulatory properties.

Gammaproteobacteria get energy for their growth from different carbon sources using either glycolysis or alternative metabolic pathways induced in stress conditions. These metabolic switches are coordinated by complex interplay of regulatory proteins sensing concentrations of available metabolites by mechanisms yet to be understood. Here, we use two transcriptional regulators, ExuR and UxuR, controlling d-galacturonate (d-gal) and d-glucuronate metabolism in Escherichia coli, as the targets for computational search of low-molecular compounds capable to bind their ligand-binding domains.

Visualizing the activity of Escherichia coli divergent promoters and probing their dependence on superhelical density using dual-colour fluorescent reporter vector.

Mosaic pattern of transcription in alternating directions is a common feature of prokaryotic and eukaryotic genomes which rationality and origin remain enigmatic. In Escherichia coli approximately 25% of genes comprise pairs of topologically linked divergently transcribed units. Given that transcriptional complex formation at each promoter in the pair induces topological changes and is itself sensitive to DNA structural perturbations, study of the functional anatomy in such areas requires special approaches.

Modes of Escherichia coli Dps Interaction with DNA as Revealed by Atomic Force Microscopy.

Multifunctional protein Dps plays an important role in iron assimilation and a crucial role in bacterial genome packaging. Its monomers form dodecameric spherical particles accumulating ~400 molecules of oxidized iron ions within the protein cavity and applying a flexible N-terminal ends of each subunit for interaction with DNA. Deposition of iron is a well-studied process by which cells remove toxic Fe2+ ions from the genetic material and store them in an easily accessible form.

Promoter islands as a platform for interaction with nucleoid proteins and transcription factors.

Seventy-eight promoter islands with an extraordinarily high density of potential promoters have been recently found in the genome of Escherichia coli. It has been shown that RNA polymerase binds internal promoters of these islands and produces short oligonucleotides, while the synthesis of normal mRNAs is suppressed. This quenching may be biologically relevant, as most islands are associated with foreign genes, which expression may deplete cellular resources.

Promoters of Escherichia coli versus promoter islands: function and structure comparison.

Expression of bacterial genes takes place under the control of RNA polymerase with exchangeable σ-subunits and multiple transcription factors. A typical promoter region contains one or several overlapping promoters. In the latter case promoters have the same or different σ-specificity and are often subjected to different regulatory stimuli.

High-density transcriptional initiation signals underline genomic islands in bacteria.

Genomic islands (GIs), frequently associated with the pathogenicity of bacteria and having a substantial influence on bacterial evolution, are groups of "alien" elements which probably undergo special temporal-spatial regulation in the host genome. Are there particular hallmark transcriptional signals for these "exotic" regions? We here explore the potential transcriptional signals that underline the GIs beyond the conventional views on basic sequence composition, such as codon usage and GC property bias. It showed that there is a significant enrichment of the transcription start positions (TSPs) in the GI regions compared to the whole genome of Salmonella enterica and Escherichia coli.

Comparative genomic analysis of the hexuronate metabolism genes and their regulation in gammaproteobacteria.

The hexuronate metabolism in Escherichia coli is regulated by two related transcription factors from the FadR subfamily of the GntR family, UxuR and ExuR. UxuR controls the d-glucuronate metabolism, while ExuR represses genes involved in the metabolism of all hexuronates. We use a comparative genomics approach to reconstruct the hexuronate metabolic pathways and transcriptional regulons in gammaproteobacteria.