Project description:We wanted to identify genes regulated by the two-component signal transduction system RR/HK06 in Streptococcus pneumoniae. We used microarray in order to discover regulated genes, investigating the effects of the over-expression of RR06 in D39 (serotype 2). Keywords: Gene Regulation
Project description:We wanted to identify genes regulated by the two-component signal transduction system RR/HK06 in Streptococcus pneumoniae. We used microarray in order to discover regulated genes, investigating the effects of the over-expression of RR06 in TIGR4 (serotype 4). Keywords: Gene Regulation
Project description:Pneumococcal bacteriocins (pneumocins) are antibacterial toxins that mediate intra- and interspecies competition within the human host. What triggers pneumocin expression is poorly understood. Using RNA-sequencing, we mapped the regulon of the pneumocin cluster (blp) of Streptococcus pneumoniae D39 and show that several antibiotics activate the blp-genes. Real-time gene expression measurements showed that while the promoter driving expression of the two-component regulatory system blpS/H was constitutive, the remaining blp-promoters (e.g. controlling pneumocin expression, immunity and the inducer peptide BlpC) was pH-dependent, induced in the late exponential phase and occurred in all cells. Intriguingly, competence for genetic transformation, mediated by the ComD/E two-component quorum system, was induced by the same stimuli. To test for regulatory interplay, we utilized synthetic BlpC and competence-stimulating peptide (CSP). Strikingly, immediately upon addition of CSP, the blp-promoters were activated in a comD/E dependent manner. After a delay, blp-expression was highly induced but strictly dependent on the presence of blpRH and blpC. This raised the question how BlpC is exported since phylogenetic analysis showed that the putative exporter for BlpC, blpAB, is not intact in strain D39 and most other strains. However, all sequenced pneumococcal strains contain intact comAB genes, encoding the transport system for CSP. In fact, we show that high expression of the blp-genes requires comAB. Together, we demonstrate that regulation of pneumocin expression is intertwined with competence, explaining why certain antibiotics induce blp-expression. Antibiotic-induced pneumocin expression might promote survival under antibiotic stress by killing and liberating DNA from neighboring (antibiotic-resistant) bacteria, which can then be used for transformation or repair.
Project description:Pneumococcal bacteriocins (pneumocins) are antibacterial toxins that mediate intra- and interspecies competition within the human host. What triggers pneumocin expression is poorly understood. Using RNA-sequencing, we mapped the regulon of the pneumocin cluster (blp) of Streptococcus pneumoniae D39 and show that several antibiotics activate the blp-genes. Real-time gene expression measurements showed that while the promoter driving expression of the two-component regulatory system blpS/H was constitutive, the remaining blp-promoters (e.g. controlling pneumocin expression, immunity and the inducer peptide BlpC) was pH-dependent, induced in the late exponential phase and occurred in all cells. Intriguingly, competence for genetic transformation, mediated by the ComD/E two-component quorum system, was induced by the same stimuli. To test for regulatory interplay, we utilized synthetic BlpC and competence-stimulating peptide (CSP). Strikingly, immediately upon addition of CSP, the blp-promoters were activated in a comD/E dependent manner. After a delay, blp-expression was highly induced but strictly dependent on the presence of blpRH and blpC. This raised the question how BlpC is exported since phylogenetic analysis showed that the putative exporter for BlpC, blpAB, is not intact in strain D39 and most other strains. However, all sequenced pneumococcal strains contain intact comAB genes, encoding the transport system for CSP. In fact, we show that high expression of the blp-genes requires comAB. Together, we demonstrate that regulation of pneumocin expression is intertwined with competence, explaining why certain antibiotics induce blp-expression. Antibiotic-induced pneumocin expression might promote survival under antibiotic stress by killing and liberating DNA from neighboring (antibiotic-resistant) bacteria, which can then be used for transformation or repair. Pairwise comparison of untreated and antibiotic-treated cells (either DNA-seq or RNA-seq). It was performed with deep sequencing, using an Illumina HiSeq 2000 machine with 100 nt paired-end reads. Control and HPUra-treated samples were analysed from duplicate samples, while other conditions were from single samples.
Project description:Streptococcus pneumoniae D39 AdcR (adhesion competence repressor) is the first metal-sensing member of the MarR (multiple antibiotic resistance repressor) family to be characterized. Expression profiling of a ∆adcR strain grown in liquid culture under microaerobic conditions revealed that transcript amounts for 13 genes were up-regulated relative to the wild-type strain, among them adcR, adcCBA, encoding a high affinity ABC uptake system for zinc, and genes encoding cell-surface zinc-binding pneumococcal histidine triad (pht) and adcII (lmb, laminin binding) proteins. Down-regulated transcripts included those encoding two putative zinc-containing alcohol dehydrogenases.
Project description:By comparing the transcriptome of Streptococcus pneumoniae wild-type D39 (serotype 2) to an isogenic strain overexpressing the response regulator of Two-Component System 7 (SPD_0158), we identified the regulon of the two-component system. In total 20 and 24 genes were considered up- and downregulated, respectively. 11 of the upregulated genes encode for proteins involved in host-glycan metabolism, while 7 encode proteins involved in carbohydrate metabolism. Most downregulated genes encode competence genes, but later experiments proved this was likely due to an atefact of overexpression. The remaining genes encode genes with other functions.
Project description:Streptococcus pneumoniae is opportunistic bacteria cause’s acute otitis media (AOM) in children. It colonizes the nasopharynx in the form of biofilms, and these biofilms act as reservoir, and are vital for pneumococcal infections. The pneumococcal biofilms are regulated by LuxS/AI-2 media quorum sensing. In this study, we confirmed the role of LuxS/AI-2 for in vitro formation of biofilms, assessed the effects of the absence of LuxS/AI-2 signaling, for pneumococcal middle ear infection and identified global genes regulated by LuxS/AI-2 during formation of pneumococcal biofilms. In the cDNA-microarray analysis, 117 genes were differentially expressed in D39 luxS mutant when compared with D39 wild type. Among the 66 genes encoding putative proteins and previously characterized proteins, 60 were significantly down-regulated and 6 were significantly up-regulated. The functional annotation revealed that genes involve in DNA replication and repair, ATP synthesis, capsule biosynthesis, cell division and cell cycle, signal transduction, transcription regulation, competence, virulence, and carbohydrate metabolism were down-regulated in the absence of LuxS/AI-2.
Project description:Streptococcus pneumoniae (pneumococcus) is a leading human respiratory pathogen that causes a variety of serious mucosal and invasive diseases. D39 is an historically important serotype 2 strain that was used in experiments by Avery and coworkers to demonstrate that DNA is the genetic material. Although isolated nearly a century ago, D39 remains extremely virulent in murine infection models and is perhaps the strain used most frequently in current studies of pneumococcal pathogenicity. To date, the complete genome sequences have been reported for only two S. pneumoniae strains; TIGR4, a recent serotype 4 clinical isolate, and laboratory strain R6, an avirulent, unencapsulated derivative of strain D39. We report herein the genome sequences of two different isolates of strain D39 and the corrected sequence and updated annotation of strain R6. Comparisons of these three related sequences allowed deduction of the likely sequence of the D39 progenitor and mutations that arose in each isolate. Despite its numerous repeated sequences and IS elements, the serotype 2 genome has remained remarkably stable during cultivation, and one of the D39 isolates contains only 5 relatively minor mutations compared to the deduced D39 progenitor. In contrast, laboratory strain R6 contains 71 single base pair changes, 6 deletions, 4 insertions, and has lost the cryptic pDP1 plasmid compared to the D39 progenitor strain. Many of these mutations are in or affect the expression of genes that play important roles in regulation, metabolism, and virulence. The nature of the mutations that arose spontaneously in these three strains, relative global transcription patterns determined by microarray analyses, and the implications of the D39 genome sequences to studies of pneumococcal physiology and pathogenesis are presented and discussed. Keywords: bacterial strain comparison, bacterial isolate comparison