GdpS contributes to Staphylococcus aureus biofilm formation by regulation of eDNA release
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ABSTRACT:
ORGANISM(S): Staphylococcus aureus
PROVIDER: GSE48656 | GEO | 2014/06/30
SECONDARY ACCESSION(S): PRJNA210961
REPOSITORIES: GEO
Similar Datasets
Project description:In Staphylococcus aureus, the role of the GGDEF domain containing protein GdpS remains poorly understood. Previous studies reported that gdpS mutant strains had decreased biofilm formation due to changes in icaADBC expression that were independent of cyclic-di-GMP levels. We deleted gdpS in three unrelated S. aureus isolates, and analyzed the resultant mutants for alterations in biofilm formation, metabolism and transcription. Dynamic imaging during biofilm development showed that GdpS inhibited early biofilm formation in only two out of the three strains examined, without affecting bacterial survival. However, quantification of biofilm formation using crystal violet staining revealed that inactivation of gdpS affected biofilm formation in all three studied strains. Extraction of metabolites from S. aureus cells confirmed the absence of cyclic-di-GMP, suggesting that biofilm formation in this species differs from that in other Gram-positive organisms. In addition, targeted mutagenesis demonstrated that the GGDEF domain was not required for GdpS activity. Transcriptomic analysis revealed that the vast majority of GGDEF-regulated genes were involved in virulence, metabolism, cell wall biogenesis and eDNA release. Finally, expression of lrgAB or deletion of cidABC in a strain lacking gdpS confirmed the role of GdpS on regulation of eDNA production that occurred without an increase in cell autolysis. In summary, S. aureus GdpS contributes to cell-to-cell interactions during early biofilm formation by influencing expression of lrgAB and cidABC mediated eDNA release. We conclude that GdpS acts as a negative regulator of eDNA release. Three strains UAMS-1, SA113 and SA564 were used in this study to compare wt with gdpS mutant after 5 hours of growth in static conditions (biofilm formation).
2014-06-30 | E-GEOD-48656 | biostudies-arrayexpress
Project description:<p>Cyclic di-GMP (c-di-GMP) is a well-known second messenger that plays a key role in many physiological processes in bacteria. The synthesis of lipids is essential for bacterial biofilm formation. However, whether c-di-GMP signaling modulates the synthesis of lipid and further regulates biofilm formation in mycobacteria is unclear, and the c-di-GMP receptor involved remains unknown. In this study, we characterized the nucleoid-associated protein (NAP) Lsr2 as a novel c-di-GMP receptor in mycobacteria. c-di-GMP specifically binds to Lsr2 at a ratio of 1:1. We showed that c-di-GMP promotes mycobacterial biofilm formation in a manner dependent on Lsr2. Furthermore, Lsr2 mediates the synthesis of keto-mycolic acid, the lipid component of the mycobacterial cell wall, by positively regulating the expression of HadD, a (3R)-hydroxyacyl-ACP dehydratase, thus, Lsr2 ultimately controls biofilm formation. Finally, c-di-GMP promotes the positive regulation of HadD by Lsr2 and mycobacterial biofilm formation. Thus, we report a novel c-di-GMP receptor that links the second messenger’s function to lipid synthesis and biofilm formation in mycobacteria.</p>
2023-12-12 | MTBLS7053 | MetaboLights
Project description:Bacterial lifestyles are dictated by the conditions encountered during colonization of a specific ecological niche or host. A wide range of environmental stimuli triggers sensory modules, that modify activity of proteins, or/and initiate a transfer of information in complex regulatory networks, which modulate gene expression, and therefore adaptation. The transition between planktonic and biofilm growth is dependent on the homeostasis of the intracellular second messenger c-di-GMP. High c-di-GMP levels driven by diguanylate cyclase (DGC) activity favor biofilm while low levels maintained by phosphodiesterase (PDE) enzymes encourage planktonic style or biofilm dispersal. In Pseudomonas aeruginosa over 40 PDE/DGC are involved in c-di-GMP homeostasis, including 16 dual enzymes possessing both canonical DGC and PDE motifs, i.e. GGDEF and EAL, respectively. It has been previously reported that the deletion of the PDE/DGC dual enzyme PA0285, one of 5 c-di-GMP-related enzymes conserved across all Pseudomonas species, impacts biofilms and causes elevated c-di-GMP levels in P. aeruginosa. Here we confirm that this role is conserved in various P. aeruginosa (PAO1, PA14 and PAK) and Pseudomonas putida strains. Deletion of PA0285 has the highest impact during the early stage of surface colonization, and RNA-seq analysis in biofilm context indicates that it may be associated with upregulation of cupA fimbrial genes. We demonstrate that the C-terminal portion of PA0285 encompassing the GGDEF and EAL domains binds the respective GTP (GGDEF) and c-di-GMP (EAL) substrates but only exhibits PDE activity in vitro. Both GGDEF and EAL domains are important for PA0285 PDE activity in vivo as suggested by site-directed mutants in each of the key motifs. Complementation of the PA0285 mutant strain with the C-terminal GGDEF/EAL portion in trans is not as effective as with the full-length gene suggesting that the transmembrane region and PAS domains contained in PA0285’s N-terminal portion influence its PDE activity in vivo. We speculate that the transmembrane portion or/and PAS domains of PA0285 impede its PDE activity in vivo through modulating the conformation of the enzyme, as shown for P. aeruginosa RbdA, in response to an unknown stimulus.
2023-07-01 | GSE223663 | GEO
Project description:Shewanella spp. possess a broad respiratory versatility, which contributes to the occupation of hypoxic/anoxic environmental or host-associated niches. Here we observed a strain-specific induction of biofilm formation in response to supplementation with the anaerobic electron acceptors dimethyl sulfoxide (DMSO) and nitrate in a panel of Shewanella algae isolates. The respiration-driven biofilm response is not observed in DMSO and nitrate reductase deletion mutants of the type strain S. algae CECT 5071, and can be restored upon complementation with the corresponding reductase operon(s) but not by an operon containing a catalytically inactive nitrate reductase. The distinct transcriptional changes, proportional to the effect of these compounds on biofilm formation, include cyclic di-GMP (c-di-GMP) turnover genes. In support, ectopic expression of the c-di-GMP phosphodiesterase YhjH of Salmonella Typhimurium but not its catalytically inactive variant decreased biofilm formation. The respiration-dependent biofilm response of S. algae may permit differential colonization of environmental or host niches.
2020-12-05 | GSE133627 | GEO
Project description:The cholera disease bacterium V. cholerae, can adopt planktonic or biofilm lifestyles depending on the intracellular concentration of the second messenger cyclic diguanylic acid (c-di-GMP). Biofilm formation protects Vibrios from stressful conditions and facilitates disease transmission by enhancing infectivity. The histone-like nucleoid structuring protein (H-NS) is a global regulator of genes associated with pathogenicity and responses to environmental stresses. H-NS represses the transcription of genes vpsT, vpsA and vpsL, which are required for the biosynthesis of the biofilm exopolysacchide matrix. Here we demonstrate that the c-di-GMP-binding protein VpsT disrupts H-NS nucleoprotein complexes at the vpsA and vpsL promoters and that this effect is enhanced by c-di-GMP. We used ChIP coupled with Next Generation Sequencing (ChIP-Seq) and transcriptome analysis (RNA-Seq) to identify additional loci repressed by H-NS affecting biofilm formation. This study showed that H-NS directly represses the transcription of genes encoding proteins present in the biofilm matrix such as the rbmA-F cluster, hemolysin and chitinase. Similar to vpsA and vpsL, the promoter region of vpsU, rbmA and rbmF exhibited overlapping H-NS and VpsT binding motifs. Deletion of vpsT increased H-NS occupancy at the vpsU, vpsA, vpsL, rbmA and rbmF promoters. Conversely, artificially increasing the c-di-GMP pool diminished H-NS occupancy at the above promoters. Deletion of vpsT did not affect H-NS occupancy at its own promoter. However, deletion of genes encoding the regulators AphA and VpsR significantly increased H-NS occupancy at the vpsT promoter. In sum, our study shows that c-di-GMP enhances biofilm formation by acting through VpsT to activate an H-NS anti-repression cascade. The Binding profile of V. cholerae H-NS to the genome was determined by ChIP followed by Next Generation Sequencing (ChIP-Seq) using the Illumina HiSeq2000 platform. V. cholerae C7258 cells expressing H-NS-FLAG fusion protein from the hns transcription and translation signals were collected from LB cultures grown to mid-exponential phase (OD600 0.5). An anti-FLAG Immunoprecipitation (IP) and an Input samples were used for the analysis.
2015-05-13 | E-GEOD-64249 | biostudies-arrayexpress
Project description:The cholera disease bacterium V. cholerae, can adopt planktonic or biofilm lifestyles depending on the intracellular concentration of the second messenger cyclic diguanylic acid (c-di-GMP). Biofilm formation protects Vibrios from stressful conditions and facilitates disease transmission by enhancing infectivity. The histone-like nucleoid structuring protein (H-NS) is a global regulator of genes associated with pathogenicity and responses to environmental stresses. H-NS represses the transcription of genes vpsT, vpsA and vpsL, which are required for the biosynthesis of the biofilm exopolysacchide matrix. Here we demonstrate that the c-di-GMP-binding protein VpsT disrupts H-NS nucleoprotein complexes at the vpsA and vpsL promoters and that this effect is enhanced by c-di-GMP. We used ChIP coupled with Next Generation Sequencing (ChIP-Seq) and transcriptome analysis (RNA-Seq) to identify additional loci repressed by H-NS affecting biofilm formation. This study showed that H-NS directly represses the transcription of genes encoding proteins present in the biofilm matrix such as the rbmA-F cluster, hemolysin and chitinase. Similar to vpsA and vpsL, the promoter region of vpsU, rbmA and rbmF exhibited overlapping H-NS and VpsT binding motifs. Deletion of vpsT increased H-NS occupancy at the vpsU, vpsA, vpsL, rbmA and rbmF promoters. Conversely, artificially increasing the c-di-GMP pool diminished H-NS occupancy at the above promoters. Deletion of vpsT did not affect H-NS occupancy at its own promoter. However, deletion of genes encoding the regulators AphA and VpsR significantly increased H-NS occupancy at the vpsT promoter. In sum, our study shows that c-di-GMP enhances biofilm formation by acting through VpsT to activate an H-NS anti-repression cascade.
2015-05-13 | GSE64249 | GEO
Project description:Background: Staphylococcus epidermidis (SE) has emerged as one of the most important causes of nosocomial infections. The SaeRS two-component signal transduction system (TCS) influences virulence and biofilm formation in Staphylococcus aureus. The deletion of saeR in S. epidermidis results in impaired anaerobic growth and decreased nitrate utilization. However, the regulatory function of SaeRS on biofilm formation and autolysis in S. epidermidis remains unclear. Results: The saeRS genes of SE1457 were deleted by homologous recombination. The saeRS deletion mutant, SE1457DsaeRS, exhibited increased biofilm formation that was disturbed more severely (a 4-fold reduction) by DNase I treatment compared to SE1457 and the complementation strain SE1457saec. Compared to SE1457 and SE1457saec, SE1457DsaeRS showed increased Triton X-100-induced autolysis (approximately 3-fold) and decreased cell viability in planktonic/biofilm states; further, SE1457DsaeRS also released more extracellular DNA (eDNA) in the biofilms. Correlated with the increased autolysis phenotype, the transcription of autolysis-related genes, such as atlE and aae, was increased in SE1457DsaeRS. Whereas the expression of accumulation-associated protein was up-regulated by 1.8-fold in 1457DsaeRS, the expression of an N-acetylglucosaminyl transferase enzyme (encoded by icaA) critical for polysaccharide intercellular adhesion (PIA) synthesis was not affected by the deletion of saeRS. Conclusions: Deletion of saeRS in S. epidermidis resulted in an increase in biofilm-forming ability, which was associated with increased eDNA release and up-regulated Aap expression. The increased eDNA release from SE1457DsaeRS was associated with increased bacterial autolysis and decreased bacterial cell viability in the planktonic/biofilm states. Comparision between SE1457 wild type strain and SA1457 SaeRS mutant strain after 4 hours and 12 hours of growth
2012-01-12 | E-GEOD-29309 | biostudies-arrayexpress
Project description:Biofilms are surface-adhered bacterial communities encased in an extracellular matrix composed of polysaccharides, proteins, and extracelluar (e)DNA, with eDNA being required for the formation and integrity of biofilms. Here we demonstrate that the spatial and temporal release of eDNA is regulated by BfmR, a regulator essential for Pseudomonas aeruginosa biofilm development. The expression of bfmR coincided with localized cell death and DNA release, with high eDNA concentrations localized to the outer part of microcolonies in the form of a ring and as a cap on small clusters. Additionally, eDNA release and cell lysis increased significantly following bfmR inactivation. Genome-wide transcriptional profiling indicated that bfmR was required for repression of genes associated with bacteriophage assembly and bacteriophage-mediated lysis. In order to determine which of these genes were directly regulated by BfmR, we utilized chromatin immunoprecipitation (ChIP) analysis to identify the promoter of PA0691, termed here phdA, encoding a previously undescribed homologue of the prevent-host-death (Phd) family of proteins. Lack of phdA expression coincided with impaired biofilm development, increased cell death and bacteriophage release, a phenotype comparable to ΔbfmR. Expression of phdA in ΔbfmR biofilms restored eDNA release, cell lysis, release of bacteriophages, and biofilm formation to wild type levels. Moreover, overexpression of phdA rendered P. aeruginosa resistant to lysis mediated by superinfective bacteriophage Pf4 which was only detected in biofilms. The expression of bfmR was stimulated by conditions resulting in membrane perturbation and cell lysis. Thus, we propose that BfmR regulates biofilm development by controlling bacteriophage-mediated lysis and thus, cell death and eDNA release, via PhdA.
2011-06-01 | GSE28953 | GEO
Project description:Background: Staphylococcus epidermidis (SE) has emerged as one of the most important causes of nosocomial infections. The SaeRS two-component signal transduction system (TCS) influences virulence and biofilm formation in Staphylococcus aureus. The deletion of saeR in S. epidermidis results in impaired anaerobic growth and decreased nitrate utilization. However, the regulatory function of SaeRS on biofilm formation and autolysis in S. epidermidis remains unclear. Results: The saeRS genes of SE1457 were deleted by homologous recombination. The saeRS deletion mutant, SE1457DsaeRS, exhibited increased biofilm formation that was disturbed more severely (a 4-fold reduction) by DNase I treatment compared to SE1457 and the complementation strain SE1457saec. Compared to SE1457 and SE1457saec, SE1457DsaeRS showed increased Triton X-100-induced autolysis (approximately 3-fold) and decreased cell viability in planktonic/biofilm states; further, SE1457DsaeRS also released more extracellular DNA (eDNA) in the biofilms. Correlated with the increased autolysis phenotype, the transcription of autolysis-related genes, such as atlE and aae, was increased in SE1457DsaeRS. Whereas the expression of accumulation-associated protein was up-regulated by 1.8-fold in 1457DsaeRS, the expression of an N-acetylglucosaminyl transferase enzyme (encoded by icaA) critical for polysaccharide intercellular adhesion (PIA) synthesis was not affected by the deletion of saeRS. Conclusions: Deletion of saeRS in S. epidermidis resulted in an increase in biofilm-forming ability, which was associated with increased eDNA release and up-regulated Aap expression. The increased eDNA release from SE1457DsaeRS was associated with increased bacterial autolysis and decreased bacterial cell viability in the planktonic/biofilm states.
2012-01-12 | GSE29309 | GEO
Project description:Although the ubiquitous bacterial secondary messenger cyclic diguanylate (c-di-GMP) plays important roles in various cellular functions including the formation of biofilm in a wide range of bacteria, its function in model plant pathogen Pseudomonas syringae is largely elusive. In order to test this in P. syringae, we overexpressed a diguanylate cyclase (YedQ) and a phosphodiesterase (YhjH) that are originally from Escherichia coli, resulting in high and low c-di-GMP levels in P. syringae, respectively. Through performing genome-wide RNA sequencing of these two strains, we found that c-di-GMP regulates (i) fliN, fliE and flhA genes, which are associated with flagellar assembly, (ii) alg8 and alg44, which are related to exopolysaccaride biosynthesis pathway, (iii) pvdE, pvdP and pvsA genes, related to siderophore biosynthesis pathway, and (iv) sodA, which is a superoxide dismutase. In particular, we identified five genes sensitive to elevated c-di-GMP level and constructed five luciferase-based reporters that effectively respond to intracellular level of c-di-GMP in P. syringae, which can be used to measure c-di-GMP levels in vivo in the future. Based on the RNA-seq results, phenotypic assays confirmed that c-di-GMP regulated many important biological pathways in P. syringae, such as negative regulation of type III secretion system (T3SS) and motility as well as positive regulation of EPS production, siderophore production and oxidative stress resistance. Taken together, the present study demonstrated that c-di-GMP is closely related to virulence and stress response in P. syringae, suggesting that tuning its level can be a new strategy to protect plants from the attack of this pathogen in the future.
2019-03-31 | GSE120889 | GEO