Project description:In Burkholderia cenocepacia H111, the large surface protein BapA plays a crucial role in the formation of highly structured communities, known as biofilms. We have recently demonstrated that Quorum sensing (QS) is necessary for the maximal expression of bapA. In this study we identify a protein from the IclR family of transcriptional regulators that, in conjunction with QS, controls biofilm formation by affecting the expression of bapA. We present evidence that, in addition to BapA, BapR influences the expression of extracellular protease, swimming motility and has a profound impact in the abundance of persister cells, making this regulator an interesting target for persister and biofilm eradication.
Project description:B. cenocepacia is an opportunistic human pathogen that is particularly problematic for patients suffering from cystic fibrosis (CF). In the CF lung, bacteria grow to high densities within the viscous mucus that is limited in oxygen. Pseudomonas aeruginosa, the dominant pathogen in CF patients, is known to grow and survive under oxygen-limited to anaerobic conditions by using micro-oxic respiration, denitrification and fermentative pathways. In contrast, inspection of the genome sequences of available B. cenocepacia strains suggested that B. cenocepacia is an obligate aerobic and non-fermenting bacterium. In accordance with the bioinformatics analysis, we observed that B. cenocepacia H111 is able to grow with as little as 0.1% O2 but not under strictly anoxic conditions. Phenotypic analyses revealed that H111 produced larger amounts of biofilm, pellicle and proteases under micro-oxic conditions (0.5% - 5% O2, i.e. conditions that mimic those encountered in CF lung infection), and was more resistant to several antibiotics. RNA-Seq and shotgun proteomics analyses of cultures of B. cenocepacia H111 grown under micro-oxic and aerobic conditions showed up-regulation of genes involved in the synthesis of the exopolysaccharide (EPS) cepacian as well as several proteases, two isocitrate lyases and other genes potentially important for life in micro-oxia.
Project description:In this study, we have analysed the transcriptional profile of B. cenocepacia H111 upon artificially altering the global cellular c-di-GMP levels. A total of 111 genes was found to be differentially expressed, of which 96 genes were down-regulated when c-di-GMP concentrations were increased. The BDSF, AHL and c-di-GMP regulons overlap for the regulation of 24 genes. Intriguingly, at a high intracellular c-di-GMP level expression of the AHL synthase cepI was found to be more than 10-fold down-regulated. Among the exclusively c-di-GMP regulated genes were genes of the phenylacetate catabolic pathway and of the type III secretion system.
Project description:Quorum sensing in Burkholderia cenocepacia H111 involves two signalling systems that depend on different signal molecules, namely N-acyl homoserine lactones (AHLs) and the diffusible signal factor cis-2-dodecenoic acid (BDSF). Previous studies have shown that AHLs and BDSF control similar phenotypic traits, including biofilm formation, proteolytic activity and pathogenicity. In this study we mapped the BDSF stimulon by RNA-Seq and shotgun proteomics analysis. We demonstrate that a set of the identified BDSF-regulated genes or proteins are also controlled by AHLs, suggesting that the two regulons partially overlap. The detailed analysis of two mutually regulated operons, one encoding three lectins and the other one encoding the large surface protein BapA and its type I secretion machinery, revealed that both AHLs and BDSF are required for full expression, suggesting that the two signalling systems operate in parallel. In accordance with this, we show that both AHLs and BDSF are required for biofilm formation and protease production.
Project description:Quorum sensing in Burkholderia cenocepacia H111 involves two signalling systems that depend on different signal molecules, namely N-acyl homoserine lactones (AHLs) and the diffusible signal factor cis-2-dodecenoic acid (BDSF). Previous studies have shown that AHLs and BDSF control similar phenotypic traits, including biofilm formation, proteolytic activity and pathogenicity. In this study we mapped the BDSF stimulon by RNA-Seq and shotgun proteomics analysis. We demonstrate that a set of the identified BDSF-regulated genes or proteins are also controlled by AHLs, suggesting that the two regulons partially overlap. The detailed analysis of two mutually regulated operons, one encoding three lectins and the other one encoding the large surface protein BapA and its type I secretion machinery, revealed that both AHLs and BDSF are required for full expression, suggesting that the two signalling systems operate in parallel. In accordance with this, we show that both AHLs and BDSF are required for biofilm formation and protease production. Burkholderia cenocepacia BDSF stimulon was determined in complex media using Illumina RNA-Seq
Project description:In Burkholderia cenocepacia H111, the large surface protein BapA plays a crucial role in the formation of highly structured communities, known as biofilms. We have recently demonstrated that Quorum sensing (QS) is necessary for the maximal expression of bapA. In this study we identify a protein from the IclR family of transcriptional regulators that, in conjunction with QS, controls biofilm formation by affecting the expression of bapA. We present evidence that, in addition to BapA, BapR influences the expression of extracellular protease, swimming motility and has a profound impact in the abundance of persister cells, making this regulator an interesting target for persister and biofilm eradication. Identification of a new regulator BapR controlling biofilm formation
Project description:Comparative transcriptional profiling of the Burkholderia cenocepacia H111 wild type, the cepR mutant H111-R and the complemented cepR mutant H111-R (pBAH27).
Project description:Members of the genus Burkholderia are versatile bacteria capable of colonizing highly diverse environmental niches. In this study, we investigated the global response of the opportunistic pathogen Burkholderia cenocepacia H111 to nitrogen limitation at the transcript and protein expression level. In addition to a classical response to nitrogen starvation, including the activation of glutamine synthetase, PII proteins and the two component regulatory system ntrBC, B. cenocepacia H111 also up-regulated polyhydroxybutyrate (PHB) accumulation and exopolysaccharide (EPS) production in response to nitrogen shortage. A search for consensus sequences in promoter regions of nitrogen responsive genes identified a s54 consensus sequence. The mapping of the s54 regulon as well as the characterization of a s54 mutant suggests an important role of s54 not only in control of nitrogen metabolism, but also in virulence of this organism.