Bacillus subtilis
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PROVIDER: PRJNA262046 | ENA |
REPOSITORIES: ENA
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Project description:Gene expression can be highly heterogeneous in clonal cell populations. An extreme type of heterogeneity is the so-called bistable or bimodal expression, whereby a cell can differentiate into two alternative expression states, and consequently a population will be composed of cells that are ‘ON’ and cells that are ‘OFF’. Stochastic fluctuations of protein levels, also referred to as noise, provide the necessary source of heterogeneity that must be amplified by autostimulatory feedback regulation to obtain the bimodal response. A classical model of bistable differentiation is the development of genetic competence in Bacillus subtilis. Noise in expression of the transcription factor ComK ultimately determines the fraction of cells that enter the competent state. Due to its intrinsic random nature, noise is difficult to investigate. We adapted an artificial autostimulatory loop that bypasses all known ComK regulators, to screen for possible factors that affect noise in the bimodal regulation of ComK. This led to the discovery of Kre, a novel factor that controls the bimodal expression of ComK. Kre appears to affect the stability of comK mRNA. Interestingly, ComK itself represses the expression of kre, adding a new double negative feedback loop to the intricate ComK regulation circuit. Our data emphasize that mRNA stability is an important factor in bimodal regulation.
2017-02-10 | GSE61757 | GEO
Project description:To cope with sudden changes in their environment, bacteria can use a bet-hedging strategy by dividing the population into cells with different properties. This so called bimodal or bistable cellular differentiation is generally controlled by positive feedback regulation of transcriptional activators. Due to continued turnover of the cytoplasmic content, it is difficult for these activators to reach an activation threshold concentration when cells are growing exponentially. This is one reason why bimodal differentiation is primarily observed from the onset of the stationary phase when exponential growth ceases. An exception is the bimodal induction of motility in Bacillus subtilis, which occurs early during exponential growth. Several mechanisms have been put forward to explain this, including double negative-feedback regulation and the stability of the mRNA molecules involved. In this study, we used fluorescence-assisted cell sorting to compare the transcriptome of motile and non-motile cells and noted that expression of ribosomal genes is lower in motile cells. This was confirmed using an unstable GFP reporter fused to the strong ribosomal rpsD promoter. We propose that the reduction in ribosomal gene expression in motile cells is the result of a diversion of cellular resources to the synthesis of the chemotaxis and motility systems. In agreement, single-cell microscopic analysis indicates that motile cells are slightly shorter than non-motile cells, an indication of slower growth. We speculate that this growth rate reduction can contribute to the bimodal induction of motility during exponential growth.
2021-01-20 | GSE164348 | GEO
Project description:In bacteria, phenotypic heterogeneity rescues the need for diversity in isogenic populations and allow concomitant multiple survival strategies when choosing only one is too risky. This powerful tactic is exploited for competence in streptococci and results in a bimodal activation, where only a subset of the community triggers the system. Deciphering the mechanisms underlying this puzzling behavior has remained challenging, especially since its study has been mainly achieved in S. mutans, where two different but interconnected regulation networks control competence. In this work, we sought to determine the origin of bimodality associated to the ComRS system thanks to the simplified S. salivarius model that harbors no supplemental signaling system. Using a single cell fluorescence reporter system together with the overexpression of the main actors of the regulation network, we showed that the ComR intracellular concentration is directly linked to the proportion of competent cells in the population. We report that this type of activation requires a functional positive feedback loop acting on comS through Opp and a putative ComS exporter, PptAB. To determine the origin of the heterogeneity amplified by the loop, we developed a mathematical model suggesting that either noise on ComS or ComR abundance could explain bimodality. Because ComR abundance is central for competence bimodal activation, we conducted an in silico identification and a systematic deletion of all the Two Component Systems (TCS) present in S. salivarius and identified CovRS, a well described virulence regulator in GAS and GBS, as a repressor of comR transcription. In vitro direct binding of CovR with the promoter of comR and transcriptomics confirmed those data. As CovRS integrates environmental stimuli that control ComR intracellular concentration, it represents the missing puzzle piece bridging environmental conditions and competence (bimodal) activation in salivarius streptococci.
2021-01-31 | GSE158512 | GEO
Project description:In this study, we have applied the top-down approach to reduce the genome of B. subtilis in order to obtain minimal strains with robust growth on complex medium at 37°C. For this purpose, we have evaluated the function of each gene of the B. subtilis genome and identified essential, important and dispensable genomic regions. Using an efficient markerless and scarless deletion method and a system allowing induction of genetic competence in the complete cell population, we have constructed two genome-reduced strains lacking about 36% of dispensable genetic information. Multi-omics analyses with the genome-reduced strains revealed substantial changes in the transcriptome, the proteome and in the metabolome. The massive reorganization of metabolism in the two genome-reduced strains can be explained by the underlying genotypes that were determined by genome re-sequencing. Moreover, the transcriptome and proteome analyses uncovered novel dispensable genomic regions that can be removed to further streamline the B. subtilis genome. In conclusion, both minimal strains show interesting metabolic features and they serve as excellent starting points to generate an ultimate reduced-genome B. subtilis cell containing only genes required for robust growth on complex medium.
2016-12-06 | PXD004565 | Pride
Project description:Here, we described comparative transcriptomic analysis of B. subtilis strain stably expressing rho with the wild type and rho-deficient strains. We show that maintaining a stable Rho level caused global changes of B. subtilis transcriptome including both strong down-regulation of the antisense transcription and considerable modifications of the sense transcription. The observed changes were more noticeable upon entering the stationary phase and comprised majority of genes controlled by global transcription regulators AbrB and CodY, competence transcription factor ComK, and stringent response. Constitutively expressed rho reprograms stationary phase-specific cellular physiology, affects adaptation of cells to nutrient limitations by attenuating the stringent response and alters cell-fate decision-making to such an extent that it blocks competence development and sporulation.
2022-02-10 | GSE195579 | GEO
Project description:Synchronizing production of antibacterial compounds and integration of DNA released by dead cells, competence is one of the most efficient bacterial evolutionary and adaptative strategies. In most streptococci, this tactic is orchestrated by the ComRS system, a pheromone communication device providing competence bimodal initiation and sharp time window of activation. Understanding how this developmental process integrates multiple inputs to fine-tune the adequate response is a long-standing question. Actually, essential genes involved in the regulation of ComRS have been challenging to study. In this work, we built a conditional mutant library using the CRISPR-interference technology and developed three complementary screens to investigate competence genetic regulation in the human commensal Streptococus salivarius. We highlighted that competence increases upon cell-wall impairment, suggesting a connection between cell envelope stress and competence activation. Notably, we report the key role played by StkP, a serine-threonine kinase known to regulate cell-wall assembly. We showed that StkP controls competence by a mechanism responding to peptidoglycan fragments. Together, those data suggest a key cell-wall sensing mechanism coupling competence to cell envelope integrity.
2022-08-25 | GSE204976 | GEO
Project description:The bacterial cell wall has been a celebrated target for antibiotics and holds real promise as a target for the discovery of new chemical matter to surmount pervasive multi-drug resistance among pathogenic bacteria. While the walls of Gram-negative bacteria are composed primarily of peptidoglycan, those of Gram-positives are more substantial and contain, in addition, large amounts of the polymer teichoic acid, covalently attached to peptidoglycan. Wall teichoic acids are a diverse group of phosphate-rich, extracellular polysaccharides that have been largely regarded as ancillary cell surface components. Recently, wall teichoic acid was shown to be essential to the proper rod-shaped cell morphology of the prototype Gram-positive bacterium Bacillus subtilis and an important virulence factor for the human pathogen Staphylococcus aureus. Thus wall teichoic acid synthesis is an intriguing target for the development of new cell wall-active antibiotics. Nevertheless, recent studies have shown that the dispensability of genes encoding teichoic acid biosynthetic enzymes in both B. subtilis and S. aureus is paradoxical and complex. Here, we report here on the discovery of a promoter (PywaC), which is sensitive to lesions in teichoic acid synthesis. Using this promoter we developed a luminescent, cell-based, reporter system to take a chemical-genetic approach to understanding the complexity of wall teichoic acid biogenesis using a large collection of antibiotics of well characterized biological activity. Our results reveal surprising interactions among undecaprenol, peptidoglycan and teichoic acid biosynthesis that help explain the complexity of teichoic acid gene dispensability. Furthermore, the new reporter assay represents an exciting avenue for the discovery of novel antibacterial molecules that impinge broadly on Gram-positive bacterial cell wall biogenesis. Keywords: comparison between depleted and repleted tagD mutant
2008-03-01 | GSE7373 | GEO
Project description:Since c-di-AMP has been implicated in the interplay of potassium and glutamate homeostasis, we analysed, using strand-specific tiling arrays (tiling step of 22 nucleotides), global gene expression in the wild type strain B. subtilis 168 at low (0.1 mM) and high (5 mM) potassium concentrations and in the presence of ammonium and glutamate as the nitrogen source as well as in the c-di-AMP free strain (∆disA, ∆cdaA, ∆cdaS) GP2222 and its isogenic suppressor mutant GP2223 that is able to grow at 5 mM potassium. In addition to strongly reduced expression of genes involved in fermentation and respiration (regulated by the NADH-responsive transcription factor Rex), genes of the SigO regulon involved in acid stress response and several competence genes was severely reduced in the mutant strain GP2222. In the wild type strain, the ktrAB and kimA genes were most strongly repressed by potassium. These genes encoding high affinity potassium uptake systems are controlled by a c-di-AMP sensitive riboswitch.Taken together, the analysis confirmed that c-di-AMP is involved in multiple cellular functions including genetic competence and revealed a particular role of genes involved in controlling NADH homeostasis.
2020-12-28 | GSE156738 | GEO
Project description:Identification of the specific WalR (YycF) binding regions on the B. subtilis chromosome during exponential and phosphate starvation growth phases. The data serves to extend the WalRK regulon in Bacillus subtilis and its role in cell wall metabolism, as well as implying a role in several other cellular processes.
2015-01-26 | GSE65272 | GEO
Project description:Expression of ykrL of Bacillus subtilis, encoding a close homologue of the Escherichia coli membrane protein quality control protease HtpX, was shown to be upregulated under membrane protein overproduction stress. Using DNA affinity chromatography, two proteins were found to bind to the promoter region of ykrL: Rok, known as a repressor of competence and genes for extracytoplasmic functions, and YkrK, a novel type of regulator encoded by the gene adjacent to ykrL, but divergently transcribed. Electrophoretic mobility shift assays showed Rok and YkrK binding to the ykrL promoter region as well as YkrK binding to the ykrK promoter region. Comparative bioinformatic analysis of the ykrL promoter regions in related Bacillus species revealed a consensus motif, which was demonstrated to be the binding site of YkrK. Deletion of rok and ykrK in a PykrL-gfp reporter strain showed that both proteins are repressors of ykrL expression. In addition, conditions which activated PykrL (membrane protein overproduction, dissipation of the membrane potential, salt- and phenol stress) point to the involvement of YkrL in membrane protein quality control. Samples for transcriptome analyses were induced at the exponential-growth phase (OD600 = 0.8) with 0.1% subtilin (subtilin containing supernatant of subtilin producing B. subtilis strain ATCC 6633). Cells were harvested 30 min after induction. Three independent cultures of each strain (target strains and controls) were used, and cells were sampled for microarray experiment. One of each strain was collected in double volume for technical replicates.
2012-04-03 | E-GEOD-33456 | biostudies-arrayexpress