Project description:UV Irradiation of the wild-type and lexA MG1655 Escherichia coli. Cells were irradiated at ~2x10^8 cells/ml in the Davis medium by 15-W germicidal lamp (254 nm, 0.66J/m^2/sec). Set also includes respective controls for unirradiated cells collected at 20 and 60 min time points. Comparison between lexA mutant and the isogenic wild-type prior the UV treatment is also included as part of the set. The SOS response in UV-irradiated Escherichia coli includes the upregulation of several dozen genes that are negatively regulated by the LexA repressor. Using DNA microarrays containing amplified DNA fragments from 95.5% of all open reading frames identified on the E. coli chromosome, we have examined the changes in gene expression following UV exposure in both wild-type cells and lexA1 mutants, which are unable to induce genes under LexA control. We report here the time courses of expression of the genes surrounding the 26 documented lexA-regulated regions on the E. coli chromosome. We observed 17 additional sites that responded in a lexA-dependent manner and a large number of genes that were upregulated in a lexA-independent manner although upregulation in this manner was generally not more than twofold. In addition, several transcripts were either downregulated or degraded following UV irradiation. These newly identified UV-responsive genes are discussed with respect to their possible roles in cellular recovery following exposure to UV irradiation. Data from the set are presented and analysed in the manuscript "Comparative gene expression profiles following UV exposure in wild type and SOS deficient Escherichia coli." Courcelle J; Khodursky A; Peter B; Brown PO; Hanawalt PC, Genetics 158: 41-64 May 2001 Keywords: other
Project description:The transcriptional changes in Escherichia coli upon induction of the SOS response are investigated by utilizing custom designed oligonucleotide microarrays. Keywords: Gene expression during the SOS response in Escherichia coli Escherichia coli K-12 MG1655 single colony in five parallells was grown to mid-log phase and exposed to UV to induce the SOS response. Total RNA was extracted from induced and uninduced cells and cDNA was prepared, fragmented and labelled prior to hybridizing to arrays. The arrays was designed to maximize the genomic coverage whilst simultaneous including only probes estimated to give an approximately uniform binding affinity. Regions coding for non-hypothetical proteins or RNAs where covered less densely than the intergenic parts.
Project description:YbjN, an enterobacteria-specific protein, is a multicopy suppressor of ts9 temperature sensitivity in Escherichia coli. Microarray study revealed that the expression level of ybjN was inversely correlated with the expression of flagellar, fimbrial and acid resistance genes. Over-expression of ybjN significantly down-regulated genes involved in the citric acid cycle, glycolysis, the glyoxylate shunt, oxidative phosphorylation, and amino acid and nucleotide metabolism. On the other hand, over-expression of ybjN up-regulated toxin-antitoxin modules, the SOS responsive pathway, cold shock proteins and starvation-induced transporter genes. Our results collectively suggest that YbjN may play important roles in regulating bacterial multicellular behaviors, metabolism and survival under various stress conditions in Es. coli. A total of 8 samples were analyzed: E. coli wild type strain (2 replicates); E. coli ybjN mutant strain (3 replicates); E. coli ybjN over-expression strain (3 replicates).
Project description:The transcriptional changes in Escherichia coli upon induction of the SOS response are investigated by utilizing custom designed oligonucleotide microarrays. Keywords: Gene expression during the SOS response in Escherichia coli
Project description:Transcriptional profiling of Escherichia coli K-12 comparing luxS mutant LW12 with wild type W3110 exposure to 10mM or 30mM hydrogen peroxide.
Project description:We used trimethylpsoralen intercalation to map supercoiling across the E. coli chromosome. We treated E. coli cells with trimethylpsoralen and exposed them to UV light. Psoralen enters cells, intercalates between DNA base pairs, and crosslinks the two strands of DNA at a rate proportional to the local superhelical density. We then fragmented DNA and hybridized crosslinked and non-crosslinked DNA fragments separately to tiling microarrays. In total 14 samples; 6 for wild-type in stationary phase (3 replicates x 2 samples as there are crosslinked and non-crosslinked samples for each replicate); 4 for wild-type in mid-exponential phase (2 replicates x 2 samples); 4 for the delta hupAB mutant in stationary phase (2 replicates x 2 samples)
Project description:Ribonucleotides are frequently incorporated into DNA and can be used as a marker of DNA replication enzymology. To investigate on a genome-wide scale, how E. coli pol V accesses undamaged chromosomal DNA during the SOS response, we mapped the location of ribonucleotides incorporated by steric gate variants of pol V across the entire E. coli genome. To do so, we used strains that are deficient in ribonucleotide excision repair (DrnhB), constitutively express all SOS-regulated genes [lexA(Def)] and constitutively “activated” RecA* (recA730). The strains also harbor two steric gate variants of E. coli pol V (Y11A or F10L), or a homolog of pol V, (pol VR391-Y13A). Ribonucleotides are frequently incorporated by the pol V-Y11A and pol VR391-Y13A variants, with a preference to the lagging strand. In contrast, the pol V-F10L variant incorporates less ribonucleotides and no strand preference was observed. Sharp transitions in strand specificity are observed at replication origin (oriC), while a gradient is observed at the termination region. To activate RecA* in a recA+ strain, we treated the strains with ciprofloxacin and genome-wide mapped the location of the incorporated ribonucleotides. Again, the polV-Y11A steric gate variant, exhibited a lagging strand preference. Our data is consistent with a specific role for pol V in lagging strand DNA synthesis across the entire E. coli genome during the SOS response.
Project description:YbjN, an enterobacteria-specific protein, is a multicopy suppressor of ts9 temperature sensitivity in Escherichia coli. Microarray study revealed that the expression level of ybjN was inversely correlated with the expression of flagellar, fimbrial and acid resistance genes. Over-expression of ybjN significantly down-regulated genes involved in the citric acid cycle, glycolysis, the glyoxylate shunt, oxidative phosphorylation, and amino acid and nucleotide metabolism. On the other hand, over-expression of ybjN up-regulated toxin-antitoxin modules, the SOS responsive pathway, cold shock proteins and starvation-induced transporter genes. Our results collectively suggest that YbjN may play important roles in regulating bacterial multicellular behaviors, metabolism and survival under various stress conditions in Es. coli.