Virulence effects and signaling partners modulated by Brucella melitensis light-sensing histidine kinase
Ontology highlight
ABSTRACT:
ORGANISM(S): Brucella melitensis bv. 1 str. 16M
SUBMITTER: Gary Splitter
PROVIDER: E-GEOD-42599 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:A whole genome expression comparision between wild type B. melitensis 16M and ΔBM-LOV-HK (ΔBMEII0679) B. melitensis strains grown in broth culture to determine genes directly and indirectly regulated by BM-LOV-HK.
2013-06-15 | GSE42599 | GEO
Project description:The four-carbon sugar erythritol is an important component of B. melitensis pathogenesis. To determine the transcriptional response to erythritol, B. melitensis strain 16M was grown in the presence of either glucose or erythritol as a sole carbon source. Two control samples (glucose) and two experimental samples (erythritol) were analyzed on Nimblegen B. melitensis microarray chips.
2013-03-01 | E-GEOD-37819 | biostudies-arrayexpress
Project description:Investigation of whole genome gene expression level changes in a Brucella melitensis delta prlr mutant compared to the wild type strain. The mutants analyzed in this study are further described in A. Mirabella, R-M Yanez, R.M. Delrue, S. Uzureau, M.S. Zygmunt, A. Cloeckaert, X. De Bolle, J.J. Letesson (2012). The two component system PrlS/PrlR of Brucella melitensis is required for persistence in mice and appears to respond to ionic strength. Microbiology A six chip study using total RNA recovered from three separate wild-type cultures of Brucella melitensis 16M and three separate cultures of a prlR mutant strain. Each chip measures the expression level of 3,198 genes from Brucella melitensis 16M with nineteen 60 mer probe pairs (PM/MM) per gene, with three-fold technical redundancy.
2012-12-31 | E-GEOD-34633 | biostudies-arrayexpress
Project description:The transcriptional regulator MucR was identified as highly attenuated in a mutagenesis screen conducted by this laboratory. MucR has been described as a key regulator of exopolysaccharide synthesis in closely related Rhizobials soil symbionts. The goal of this study was to apply custom Brucella melitensis oligonucleotide microarrays to identify the regulatory targets of MucR. These studies revealed that MucR's regulation of exopolysaccharide synthesis is also conserved in Brucella melitensis and is also a key regulator of iron sequestering/storage and denitrification. Keywords: Microarray comparison of a genetically modified organism compared to wild type. There are 3 biological replicates isolated from Brucella melitensis wild type 16M and from 16M∆mucR.
2010-04-23 | E-GEOD-13647 | biostudies-arrayexpress
Project description:Brucella melitensis and Brucella canis differ by ~75 genes yet B. melitensis is highly virulent for humans while B. canis is considered rarely pathogenic. No identified bacterial factors or mechanisms account for this difference in virulence. To identify functional differences of these two bacteria, gene transcription was examined during infection of murine macrophages and compared to bacteria grown in broth. Our analysis identified transcriptional differences in macrophage infection between B. melitensis and B. canis genes involved in iron transport. Increased transcription of the TonB, enterobactin, and ferric anguibactin transport systems were observed in B. canis but not B. melitensis during infection of macrophages. Therefore, iron appears as an important requirement during the first 24h of infection by B. canis but not for B. melitensis and provides strategies for controlling these pathogens. Comparison of total bacterial RNA from Brucella melitensis infected murine macrophages to broth grown bacteria
2012-12-01 | E-GEOD-34495 | biostudies-arrayexpress
Project description:Facultative intracellular Brucella infect and survive inside macrophages, and the outcome of macrophage-Brucella interaction is a basis for establishment of a chronic Brucella infection. The majority of Brucella are killed at the early infection stage. A subpopulation of virulent Brucella strains is instead trafficked to an intracellular replicative phagosome, and are resistant to further attack and begin to multiply dramatically. Virulent Brucella also inhibit macrophage apoptosis that in turn favors pathogen survival and replication. We used the Affymetrix mouse GeneChip 430 2.0 array to analyze mouse macrophage gene expression profiles during the time course of virulent B. melitensis strain 16M infection. Experiment Overall Design: Murine J774.A1 macrophage cells were infected with B. melitensis strain 16M at a MOI of 200:1. Brucella cultures derived from different Brucella colonies were used to infect different groups of macrophage cells to reflect independent infections. Following 4 h incubation, the cells were washed with PBS and treated with 50 ug/ml gentamicin to kill extracellular Brucella. At 0 h (no Brucella infection), 4 h, 24 h, and 48 h post-infection, cells were individually collected, and total RNA was isolated. The Affymetrix mouse GeneChip 430 2.0 array was used for microarray hybridization. Microarray intensity data were obtained by using Affymetrix GCOS software and further analyzed by GeneSpring and other software programs.
2008-06-13 | E-GEOD-5202 | biostudies-arrayexpress
Project description:A mini-microarray containing spotted oligos corresponding to 289 B. melitensis genes was used to identify candidate genes whose transcription is influenced by BlxR. This microarray contains genes encoding the type IV secretion system, flagella, transcriptional regulators, transporters, and proteins involved in outer membrane biogenesis, iron acquisition, and denitrification. RNA was prepared from both B. melitensis 16M, and the blxR deletion strain, 16M(delta)blxR, grown to late log phase in TSB. Keywords: genetic modification Bacteria were grown in TSB until mid/late-log phase. Before RNA isolation, phenol-ethanol (1:19) was added, cells were pelleted by centrifugation and stored at -80M-BM-0C. Total RNA was extracted from cell pellets using the MasterPure RNA Purification Kit (Epicentre), and DNA was removed using TURBO DNA-free (Ambion). cDNA was synthesized from 7 ug of total RNA in the presence of CyDye-3-dCTP (Amersham Biosciences). The labeled cDNA was purified using the ChipShot cDNA System (Promega). For each sample, RNA was extracted from two independently grown cultures and cDNA was generated three times for each RNA sample. For the mini-microarray, 289 potential open reading frames (ORFM-bM-^@M-^Ys) of the B. melitensis 16M genome were selected based on their hypothesized involvement in virulence. These included genes for proteins involved in: type IV secretion, flagella, outer membrane biogenesis, transcriptional regulation, denitrification, iron uptake, and peptide and small molecule transport. Oligonucleotides (70-mers) representing these 289 genes were purchased from Qiagen, re-suspended in 3x SSC, and spotted onto Ultra GAPS coated slides (Corning) using the MicroGrid Microarrayer (Apogent Discoveries). Slides were desiccated in a vacuum oven with Drierite for 48 h and UV cross linked using a Stratalinker at 600 mJ. The printing of each ORF spot was verified by hybridizing with Cy3 labeled 9 mer (Qiagen). The slides were stored under vacuum at room temp with M-bM-^@M-^\DrieriteM-bM-^@M-^] pellets until further use. 16M genomic DNA was labeled by incorporation of CyDye-5-dCTP (Amersham Biosciences) using High Prime DNA Labeling (Roche) according to manufacturerM-bM-^@M-^Ys instructions. Unincorporated nucleotides were removed using Micron YM-30 Columns (Millipore). Cy3-labeled cDNA was mixed with 0.75 ug cy5-labeled gDNA, and hybridization buffer (Pronto Plus Kit, Promega) was added to a final volume of 35 ul. The entire sample was loaded onto the printed array and covered with a HybriSlip coverslip (Molecular Probes). Arrays were placed in specialized hybridization chambers (Telechem Inc., Sunnyvale, CA), and incubated at 42M-BM-0C overnight. Slides were washed using the Pronto Plus Hybridization System (Promega) and dried by centrifugation at 1000 rpm for 5 min. Slides were scanned using the GenePix4000B scanner (Axon Instruments) with independent excitation of the fluorophores Cy3 and Cy5. Background intensities for each spot were calculated using GenPixPro 3.0 software (Axon Instruments) via the segmentation method. The signal intensity for each spot was calculated as the difference between the average signal intensity and the average local background intensity. Data was normalized to gDNA levels by calculating the ratios of cDNA to gDNA for each spot. Data was then log2 transformed, and the signal ratio for each spot was divided by the mean signal ratio of the entire array to validate comparisons between different arrays. Data normalization and statistical analysis were performed using TIGR MeV. GPR files were not submitted because they cannot be located.
2008-04-30 | E-GEOD-10735 | biostudies-arrayexpress
Project description:Quorum sensing (QS) is a communication system in bacteria that regulates gene expression in response to a small diffusible signal indicative of the bacterial population present. QS has been shown to regulate virulence genes, biofilm formation, cell division and secretion systems in a diverse set of bacteria. Brucella melitensis produces a QS signaling molecule, N-dodecanoyl homoserine lactone (C12-HSL), that interacts with a LuxR transcriptional regulator, VjbR. Deletion of vjbR has been found to highly attenuate intracellular survival of B. melitensis. The importance of QS in the regulation of genes necessary for the establishment and maintenance of B. melitensis infection has already been described for the virB and fla operons, however, a complete description of the vjbR regulon has not been described. Genome mining revealed seven luxR-like genes in B. melitensis, only one of which, vjbR, was confirmed to be attenuated for intracellular survival using macrophage assays. Using custom B. melitensis microarrays, we compared transcripts from wild type and ∆vjbR strains in the presence and absence of exogenous C12-HSL. Comparison of the transcriptomes obtained from culture-grown bacteria revealed bi-phasic gene regulation with expression peaks during exponential and early stationary growth phase; characterized by the regulation of 226 and 246 genes (respectively) by VjbR and 349 and 146 (respectively) altered by the addition of C12-HSL. Comparison of the VjbR and C12-HSL regulated genes provided confirmation that expression of 134 genes are regulated by both conditions with all but 3 genes inversely regulated. Overall, these results indicate that VjbR is an activator of gene expression at the exponential growth phase and exerts an equal effect on gene expression at the stationary growth phase; while C12-HSL has a repressive effect on gene expression at both growth stages examined and acts as an antagonist to VjbR activity. Direct transcript analysis comparing ∆vjbR with and without the addition of C12-HSL revealed that the AHL signal is able to direct gene expression in the absence of VjbR and exclusively exerted a positive influence on the expression of 56 genes, including a 93-fold increase in the expression of BabR, a second LuxR homologue. Genes regulated by these QS components included adhesins, proteases, antibiotic and toxin resistance genes, stress survival aids, including DNA repair genes and protein chaperones, transporters and porins, cellular membrane biogenesis genes, amino acid metabolism and transport, transcriptional regulators, and energy production genes. From these data, we conclude that QS is a global regulator of gene expression, and present potential virulence genes that may provide insight into the bacteria’s ability to establish and maintain the replicative vacuole (BCV) within the host cell. Keywords: Microarray comparison of a genetic deletion mutant and the addition of an effector molecule. Samples consist of RNA isolated from Brucella melitensis grown to logarithmic or stationary phase. RNA was extracted from wild type with and without exogenously added C12-HSL, and a ∆vjbR mutant. There are three biological replicates of each sample. Every Brucella melitensis open reading frame was printed in quadruplicate on each microarray. Each replicate was normalized against labeled Brucella melitensis 16M genomic DNA.
2010-04-23 | E-GEOD-13527 | biostudies-arrayexpress
Project description:Quorum sensing (QS) is a communication system in bacteria that regulates gene expression in response to a small diffusible signal indicative of the bacterial population present. QS has been shown to regulate virulence genes, biofilm formation, cell division and secretion systems in a diverse set of bacteria. Brucella melitensis produces a QS signaling molecule, N-dodecanoyl homoserine lactone (C12-HSL), that interacts with a LuxR transcriptional regulator, VjbR. Deletion of vjbR has been found to highly attenuate intracellular survival of B. melitensis. The importance of QS in the regulation of genes necessary for the establishment and maintenance of B. melitensis infection has already been described for the virB and fla operons, however, a complete description of the vjbR regulon has not been described. Genome mining revealed seven luxR-like genes in B. melitensis, only one of which, vjbR, was confirmed to be attenuated for intracellular survival using macrophage assays. Using custom B. melitensis microarrays, we compared transcripts from wild type and ∆vjbR strains in the presence and absence of exogenous C12-HSL. Comparison of the transcriptomes obtained from culture grown bacteria revealed bi-phasic gene regulation with expression peaks during exponential and early stationary growth phase; characterized by the regulation of 226 and 246 genes (respectively) by VjbR and 349 and 146 (respectively) altered by the addition of C12-HSL. Comparison of the VjbR and C12-HSL regulated genes provided confirmation that expression of 134 genes are regulated by both conditions with all but 3 genes inversely regulated. Overall, these results indicate that VjbR is an activator of gene expression at the exponential growth phase and exerts an equal effect on gene expression at the stationary growth phase; while C12-HSL has a repressive effect on gene expression at both growth stages examined and acts as an antagonist to VjbR activity. Direct transcript analysis comparing ∆vjbR with and without the addition of C12-HSL revealed that the AHL signal is able to direct gene expression in the absence of VjbR and exclusively exerted a positive influence on the expression of 56 genes, including a 93-fold increase in the expression of BabR, a second LuxR homologue. Genes regulated by these QS components included adhesins, proteases, antibiotic and toxin resistance genes, stress survival aids, including DNA repair genes and protein chaperones, transporters and porins, cellular membrane biogenesis genes, amino acid metabolism and transport, transcriptional regulators, and energy production genes. From these data, we conclude that QS is a global regulator of gene expression, and present potential virulence genes that may provide insight into the bacteria’s ability to establish and maintain the replicative vacuole (BCV) within the host cell. Keywords: Microarray comparison of a genetic deletion mutant and the addition of an effector molecule. Samples consist of RNA isolated from Brucella melitensis grown to logarithmic or stationary phase. RNA was extracted from a ∆vjbR mutant with and without exogenously added C12-HSL. There are three biological replicates of each sample. Every Brucella melitensis open reading frame was printed in quadruplicate on each microarray. Each replicate was normalized against labeled Brucella melitensis 16M genomic DNA.
2010-04-23 | E-GEOD-13633 | biostudies-arrayexpress
Project description:We sequenced the mRNA of Brucella melitensis 16M and otpR mutant under acid stress to identify genes that were regulated by regulator OtpR. Examination of mRNA levels in individual sample under acid stress.
2015-08-11 | E-GEOD-48165 | biostudies-arrayexpress