Impact of suppression of the SOS response on protein expression in clinical isolates of Escherichia coli under antimicrobial pressure of ciprofloxacin
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive HF
ORGANISM(S): Escherichia Coli
SUBMITTER: 
Benno Kuropka
LAB HEAD: Jose Manuel
PROVIDER: PXD050358 | Pride | 2024-04-04
REPOSITORIES: Pride
Dataset's files
Items per page: 5 1 - 5 of 79 |
Similar Datasets
Project description:The HIVEP/ZAS/Schnurri genes encode large zinc finger proteins that regulate gene expression through DNA binding to the kappaB motif. The ZAS proteins have also been shown to associate with signaling molecules to regulate the TNFa and TGFb signaling pathways. Because ZAS3 transcript and protein expression is rapidly abrogated in primary lymphocytes upon tissue culture, we have generated a ZAS3-null mouse model to study the physiological function of ZAS3. Mice with targeted disruption of ZAS3 are viable with life span comparable to controls. Additionally, the gross anatomy and histology of the major organs, proliferation rates of splenocytes and thymocytes, and the diversity of the TCRb chains of ZAS3 mice are comparable to wild-type. However, compared to wild-type mice, there are decreased CD3+CD4+ and CD3+CD4+CD69+ thymocyte populations. Additionally, CD44hi/CD62Llo subset and CD25 and CD69 expression are increased in splenocytes of ZAS3 mice. Microarray analysis validated by real time PCR revealed changes in the expression level of specific transcripts in ZAS3-null thymus, including several proteins in the G-protein coupled olfactory receptor family. Furthermore, EMSA shows that disruption of ZAS3 results in varying changes in binding activities towards NF-kB and AP1 binding sites. Other phenotypes observed in the ZAS3 mice include generalized increased bone density in older animals and infertility in females. As with ZAS2/shn-2 mice, disruption of ZAS3 is not lethal and does not grossly affect development or homeostasis. We propose that the ZAS proteins likely have overlapping functions, which may be uncovered with deletion of multiple ZAS genes in a single animal. Experiment Overall Design: Five independent microarray hybridizations were performed with thymocytes isolated from sex- and age-matched mutant and wild-type littermates at 6-8 weeks of age. The Agilent Whole Mouse Genome Oligo Microarray (G4121A) containing over 20,000 60-mer oligonucleotide probes representing mouse genes, ESTs and EST clusters was used (Agilent, Cincinnati, OH). Probe labeling and hybridization were preformed by the Microarray Core Faculty (Columbus Childrenâs Research Institute) following the manufacturer's protocols. Briefly, cDNA probes were synthesized with Superscript III, oligo-dT primers and dNTPs supplemented with amino-allyl-UTP to improve hybridization characteristics and stability. After purification, cDNA samples from mutant or control mice were labeled with Cy3 or Cy5, respectively, and hybridized to the microarray for 14 hours at 48o C. Slide image was acquired using an Affymetrix 428 scanner with gain settings set so that 95% of spots were below saturation to yield the maximum dynamic range within an experiment. Images were converted into â.gprâ files using GenePix software (Axon, Union City, CA). Data were analyzed using GeneTraffic 2.6 software (Iobion, La Jolla, CA). Lowess-global normalization was applied to all experiments. Flagging parameters were set as spot intensity lower than the intensity of local spot background, spot intensity lower than average background, and raw spot intensity less than 100. Flagged spots were not included in normalization or aggregate calculations. Only genes that were induced or repressed 1.5-fold in at least four of the five independent analyses and hybridization were targeted for potential further study.
2008-06-14 | E-GEOD-6570 | biostudies-arrayexpress
Project description:Integrating laterally acquired virulence genes into the backbone regulatory network is important for the pathogenesis of Escherichia coli O157:H7, which has captured many virulence genes through horizontal transfer during evolution. GadE is an essential transcriptional activator of glutamate decarboxylase (GAD) system, the most efficient acid resistance mechanism in E. coli. The full contribution of GadE to the acid resistance and virulence of pathogenic E. coli O157:H7 remains largely unknown. We inactivated gadE in E. coli O157:H7 Sakai and compared global transcription profiles with that of wild type in exponential and stationary phases of growth using microarrays containing 6088 ORFs from three E. coli genomes. gadE inactivation significantly altered the expression of 60 genes independent of growth phase and 122 genes in a growth phase-dependent manner. Inactivation of gadE markedly down-regulated the expression of gadA, gadB, gadC and many acid fitness island genes in a growth phase-dependent manner. Nineteen genes encoded on the locus of enterocyte effacement (LEE), including ler, showed a significant increase in expression upon gadE inactivation. Altogether, our data indicate that GadE is critical for acid resistance of E. coli O157:H7 and plays an important role in virulence by down-regulating expression of LEE. The results are based on O157:H7 Sakai wild type and gadE mutant exponential and stationary phase cultures grown in MOPS minimal medium. Differences in transcript levels were determined using a mixed model ANOVA in R/MAANOVA which tested for significant differences due to growth phase (exponential or stationary), strain (wild type or mutant) and the interaction of these two factors using the following linear model: array+dye+sample (biological replicate)+ phase+strain+phase*strain. We incorporated the dye-swaps among the biological replicates.
2010-05-18 | E-GEOD-13132 | biostudies-arrayexpress
Project description:Transcriptional profile of the uppS-RBS (A to C) mutant during log growth phase in LB medium. Bacillus subtilis W168, WT vs. uppS-RBS (A to C). The experiment was conducted two times using three independent total RNA preparations (biological triplicates). Each comparison was performed in dye-swap with the same RNA preparation. For each paried comparison, one sample was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647.
2013-04-01 | E-GEOD-45305 | biostudies-arrayexpress
Project description:This analysis revealed ~17.% of all ORFs (292 out of a total 1697 ORFs) differed by at least 1.5 fold (Log2 +0.6) between the mutant and wild-type strains. Of these, ~45% of the ORFs (130 out of 292 ORFs) revealed at least 2-fold (Log2 +1) differences. The complete spectrum of differentially expressed genes in the muatnt strain revealed almost equal number of genes up (143, 49%) or down-regulated (149, 51%). However, at a higher stringency (at least 2-fold), this distribution revealed slightly more leaning towards down-regulated genes (74 out of 130, 57%) indicating the lack of SP-STK results into down regulation of several genes. Amongst the genes belonging to different functional categories, the most notable genes that were affected by the absence of SP-STK were those correspond to Carbohydrate transport and metabolism (57 out of 292, 20% at low stringency and 32 out of 130 ,~25% at high stringency), phage (28, ~10% at low stringency and 24, ~18% at high stringency), and virulence (5-6% irrespective of stringency level). Almost 20-25% of differentially expressed genes revealed at two stringencies were found to be either with unknown or undefined functions. Keywords: knockout mutant This analysis is based on a total of three independent prearations of total RNA from the wild-type S. pyogenes M1SF370 and the isogenic knockout mutant lacking SP-STK (SPy1625). First two preparations included dyeswap experiment and hence repeated twice (exp1-4). The third preparation done only one (Exp#5). Thus, the present analysis is based on five separate experiments
2013-02-28 | E-GEOD-10914 | biostudies-arrayexpress
Project description:Borelia burgdorferi, the causative agent of the Lyme disease, cycles between a vector (tick) and the mammalian host. Our principal goal is to study how B. burgdorferi adapts its metabolism to colonize and survive in the tick. We established the first growth-phase related acetylome and demonstrated the role of acetylation on central metabolism regulation. We also demonstrated that acetyl phosphate is the primary acetyl donor in B. burgdorferi.
2018-09-24 | PXD010065 | Pride
Project description:The object of this study was to explore whether the loss of mRNA decapping in the Arabidopsis mutant tdt-1 resulted in global dfferences of RNA profiles, as compared to wild type. Experiment Overall Design: In this experiment we compared expression in tdt-1 and wild type 3 day whole seedlings. We performed three biological replicates, in which one experiment was dye-swapped.
2008-06-15 | E-GEOD-7359 | biostudies-arrayexpress
Project description:We investigated the genome-wide DNA binding of the chromosome partitioning and sporulation protein and ParB family member Spo0J in Bacillus subtilis using chromatin immunoprecipitation and DNA microarrays. We identified ten parS loci to which Spo0J binds, two of which were unexpectedly distant (>1 Mb) from the origin of replication. We used all ten sites to refine the consensus sequence for parS. We found that Spo0J spreads along the DNA around each site. Binding was near maximal levels up to 1.6 kb away from parS, and significantly above background as far away as 18 kb. Deletion of soj (parA) had no detectable effect on spreading. In contrast, the spo0J93 allele appeared to cause a significant decrease in spreading in vivo, without significantly affecting the DNA binding affinity in vitro. spo0J93 causes a phenotype similar to that of a spo0J null mutant and alters the region thought to be involved in interaction between Spo0J dimers. Our findings indicate that spreading is important for in vivo function of Spo0J. Gene expression in areas near parS sites was similar in wild-type and a spo0J null mutant, indicating that binding and spreading of Spo0J on DNA does not silence transcription of nearby genes. Keywords: genetic modification Gene expression was compared in spo0J mutant and wild-type B. subtilis by isolating total RNA from three independent cultures of each strain and comparing them on microarrays. One of the arrays (GSM152506) used the reverse dye assignments relative to the other two.
2010-06-11 | E-GEOD-6598 | biostudies-arrayexpress
Project description:Transcriptomes from the hat1, hat2 and hat1 hat2 mutants in S288C yeast background.
2010-06-09 | E-GEOD-2434 | biostudies-arrayexpress
Project description:A total of 163 genes were found to be differentially expressed; 38 genes of them were up regulated and 125 genes were down regulated. Most notably the functional categories that were affected include Virulence genes (18 genes 11% of the total significantly differentiated genes), carbohydrate metabolism and cell envelop realted genes ( 31, 19.1% of the total), and aminoacid transport genes (21genes, 12.9%). Among the virulence-related genes, the most notable one were those belong to the production of capsule, the M protein, exotoxin, NAD glycohydrolase and Streptolysin S. Some of the capsule related genes were down regulated by more than 64 fold. These results indicate that CdhA (group A Streptococcal Cell division controlling and Chain-forming cell wall hydrolase) plays an important role in the regulation of virulence. S.pyogenes strain SF370 (wild-type) was procured from ATCC (ATCC 700294). M1-CdhA(-) mutant strain was obtained using pFW5 vector containing spectinomycin resistance marker (aad9). The mutant lacking CdhA is highly defective in cell division and growth, lacks antiphagocytic property and attenuated for virulence. Both strains (M1-Wild-type and M1-CdhA(-) were grown in Todd-Hewitt broth to their late log phase. Bacteria were harvested by centrifugation and washed twice with sterile PBS. Total RNA was isolated from these washed streptococci using Qiagen RNeasy Mini kit. For synthesis and labeling of cDNA, 20 ug of total RNA, 1.5 ug of random hexamer, 0.5mM dNTPs (except that 0.2mM of dTTP was replaced by the same amount of amino-allyl dUTP to incorporate dUTP into first-strand cDNA) were used in each reverse transcriptase reaction (Superscript II Reverse Transcriptase, Invitrogen). Purified cDNA preparations from the wild-type and M1CdhA(-) mutant strains were labeled with either Alexafluor-555 or Alexafluor-647 depending on the experimental design ( i.e. dye swap experiment). Differentially labeled probes were then combined and purified. Using three independently isolated RNA preparations (biological replicates), a total of 11 experiments (incorporating dye swaps) were performed. Accordingly eleven hybridization measurements for this mutant were performed. Thus Exp-1 to -4 (GSM388703, GSM388717, GSM388718 and GSM388719) are the technical replicates of the biological sample-1, Exp-5 to 8(GSM388732, GSM388733, GSM388734, and GSM388735) are technical replicates of biological sample-2 and Exp-9,-10, and -11 (GSM388736,GSM388737, and GSM388738) are technical replicates of the biological sample-3. Signals of the bound reagents on the microarray spots in terms of relative fluorescence values were measured and quantified by a laser scanner (GenePix 4100 ) at 10um/pixel resolution. The resulting images were processed using Gene Pix Pro software (version 4.0, Axon instruments). The raw data were obtained in the form of GenPix *.gpr output files. The web application CARMAweb (Comprehensive R based microarray analysis web service) was used for the normalization and analysis of microarray data. All raw data (in the form of *. GPR files) were uploaded to the web application in the data directory. Using appropriate navigation tree, background correction from the foreground signal was applied and within microarray normalization was achieved using the Lowess method. Genes flagged as bad spots by the scanning software were excluded from the analysis and all flagged spots were given a weight of zero. The normalized data were then subjected to fold-change analysis and t-statistics using Bioconductor multtest package. CARMAweb allows to set Log2 cut-off values for both the M (regulation) and the A (average expression) values. Differentially expressed genes (Log2 values of Mutant647-red vs. wild-type555-green or Log2 values Mutant-555(green) vs Wild-type 647-red) were defined based on cut-off value >1 Log2< i.e. all genes that show a two or more-fold up- or down-regulation. Bioconductors Multtest package provided suitable method to adjust P values according to multiple hypothesis testing problems.
2010-10-01 | E-GEOD-15546 | biostudies-arrayexpress
Project description:This series of microarrays compares gene expression by the bacterial pathogen Proteus mirabilis when the transcriptional regulator mrpJ is deleted or induced to levels found during experimental urinary tract infection. The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections. Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for successful disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects a wide array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking expression levels that occur during UTI leads to differential expression of 217 genes related to, among others, bacterial virulence, type VI secretion and metabolism. We probed the molecular mechanism of transcriptional regulation through MrpJ using reporter assays and chromatin immunoprecipitation (ChIP). Two virulence-associated target genes, the flagellar master regulator flhDC and mrp itself, appear to be regulated through a binding site proximal to the transcriptional start, complemented by a more distantly situated enhancer site. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observe that mrpJ is widely conserved in a collection of recent clinical isolates, leading us to conclude that our results elucidate an unanticipated role of MrpJ as a global regulator of P. mirabilis virulence. Four biological replicates were analyzed for each set of arrays (P. mirabilis HI4320 wild type vs. ΔmrpJ, and vector pLX3607 vs. mrpJ plasmid pLX3805).
2015-04-13 | E-GEOD-63321 | biostudies-arrayexpress