High incidence of multiple antibiotic resistant cells in cultures of in enterohemorrhagic Escherichia coli O157:H7.

The spontaneous incidence of chloramphenicol (Cam) resistant mutant bacteria is at least ten-fold higher in cultures of enterohemorrhagic Escherichia coli O157:H7 strain EDL933 than in E. coli K-12. It is at least 100-fold higher in the dam (DNA adenine methyltransferase) derivative of EDL933, compared to the dam strain of E.

High efficiency generalized transduction in Escherichia coli O157:H7.

Genetic manipulation in enterohemorrhagic E. coli O157:H7 is currently restricted to recombineering, a method that utilizes the recombination system of bacteriophage lambda, to introduce gene replacements and base changes inter alia into the genome. Bacteriophage 933W is a prophage in E.

Using stable MutS dimers and tetramers to quantitatively analyze DNA mismatch recognition and sliding clamp formation.

The process of DNA mismatch repair is initiated when MutS recognizes mismatched DNA bases and starts the repair cascade. The Escherichia coli MutS protein exists in an equilibrium between dimers and tetramers, which has compromised biophysical analysis. To uncouple these states, we have generated stable dimers and tetramers, respectively.

The alkaloid compound harmane increases the lifespan of Caenorhabditis elegans during bacterial infection, by modulating the nematode's innate immune response.

The nematode Caenorhabditis elegans has in recent years been proven to be a powerful in vivo model for testing antimicrobial compounds. We report here that the alkaloid compound Harmane (2-methyl-β-carboline) increases the lifespan of nematodes infected with a human pathogen, the Shiga toxin-producing Escherichia coli O157:H7 strain EDL933 and several other bacterial pathogens. This was shown to be unrelated to the weak antibiotic effect of Harmane.

Bleomycin sensitivity in Escherichia coli is medium-dependent.

Bleomycin (BLM) is a glycopeptide antibiotic and anti-tumor agent that targets primarily the furanose rings of DNA and in the presence of ferrous ions produces oxidative damage and DNA strand breaks. Escherichia coli cells growing in broth medium and exposed to low concentrations of BLM contain double-strand breaks and require homologous recombination to survive. To a lesser extent, the cells also require the abasic (AP) endonucleases associated with base excision repair, presumably to repair oxidative damage.

RecA-independent single-stranded DNA oligonucleotide-mediated mutagenesis.

The expression of Beta, the single-stranded annealing protein (SSAP) of bacteriophage lambda in Escherichia coli promotes high levels of oligonucleotide (oligo)-mediated mutagenesis and offers a quick way to create single or multiple base pair insertions, deletions, or substitutions in the bacterial chromosome. High rates of mutagenesis can be obtained by the use of mismatch repair (MMR)-resistant mismatches or MMR-deficient hosts, which allow for the isolation of unselected mutations. It has recently become clear that many bacteria can be mutagenized with oligos in the absence of any SSAP expression, albeit at a much lower frequency.

DNA methylation and mutator genes in Escherichia coli K-12.

Mutator strains of Escherichia coli have been used to define mechanisms that account for the high fidelity of chromosome duplication and chromosome stability. Mutant strains defective in post-replicative mismatch repair display a strong mutator phenotype consistent with a role for correction of mismatches arising from replication errors. Inactivation of the gene (dam) encoding DNA adenine methyltransferase results in a mutator phenotype consistent with a role for DNA methylation in strand discrimination during mismatch repair.

Roles of DNA adenine methylation in host-pathogen interactions: mismatch repair, transcriptional regulation, and more.

The DNA adenine methyltransferase (Dam methylase) of Gammaproteobacteria and the cell cycle-regulated methyltransferase (CcrM) methylase of Alphaproteobacteria catalyze an identical reaction (methylation of adenosine moieties using S-adenosyl-methionine as a methyl donor) at similar DNA targets (GATC and GANTC, respectively). Dam and CcrM are of independent evolutionary origin. Each may have evolved from an ancestral restriction-modification system that lost its restriction component, leaving an 'orphan' methylase devoted solely to epigenetic genome modification.

DnaC inactivation in Escherichia coli K-12 induces the SOS response and expression of nucleotide biosynthesis genes.

Background: Initiation of chromosome replication in E. coli requires the DnaA and DnaC proteins and conditionally-lethal dnaA and dnaC mutants are often used to synchronize cell populations.

Methodology/principal Findings: DNA microarrays were used to measure mRNA steady-state levels in initiation-deficient dnaA46 and dnaC2 bacteria at permissive and non-permissive temperatures and their expression profiles were compared to MG1655 wildtype cells.

YhdJ, a nonessential CcrM-like DNA methyltransferase of Escherichia coli and Salmonella enterica.

The Caulobacter crescentus DNA adenine methyltransferase CcrM and its homologs in the alpha-Proteobacteria are essential for viability. CcrM is 34% identical to the yhdJ gene products of Escherichia coli and Salmonella enterica. This study provides evidence that the E.

Hda-mediated inactivation of the DnaA protein and dnaA gene autoregulation act in concert to ensure homeostatic maintenance of the Escherichia coli chromosome.

Initiation of DNA replication in Eschericia coli requires the ATP-bound form of the DnaA protein. The conversion of DnaA-ATP to DnaA-ADP is facilitated by a complex of DnaA, Hda (homologous to DnaA), and DNA-loaded beta-clamp proteins in a process termed RIDA (regulatory inactivation of DnaA). Hda-deficient cells initiate replication at each origin mainly once per cell cycle, and the rare reinitiation events never coincide with the end of the origin sequestration period.

The beta sliding clamp binds to multiple sites within MutL and MutS.

The MutL and MutS proteins are the central components of the DNA repair machinery that corrects mismatches generated by DNA polymerases during synthesis. We find that MutL interacts directly with the beta sliding clamp, a ring-shaped dimeric protein that confers processivity to DNA polymerases by tethering them to their substrates. Interestingly, the interaction of MutL with beta only occurs in the presence of single-stranded DNA.

Dam methylation: coordinating cellular processes.

GATC sequences in Escherichia coli DNA are methylated at the adenine residue by DNA adenine methyltransferase (DamMT). These methylated residues and/or the level of DamMT can influence cellular functions such as gene transcription, DNA mismatch repair, initiation of chromosome replication and nucleoid structure. In certain bacteria, unlike E.

RecN and RecG are required for Escherichia coli survival of Bleomycin-induced damage.

The sensitivity of a panel of DNA repair-defective bacterial strains to BLM was investigated. Escherichia coli recA cells were far more sensitive than were uvrA, dam-3, and mutM mutY strains, underscoring the importance of RecA to survival. Strains recBCD and recN, which lack proteins required for double strand break (DSB) repair, were highly sensitive to BLM, while recF cells were not.

Role of SeqA and Dam in Escherichia coli gene expression: a global/microarray analysis.

High-density oligonucleotide arrays were used to monitor global transcription patterns in Escherichia coli with various levels of Dam and SeqA proteins. Cells lacking Dam methyltransferase showed a modest increase in transcription of the genes belonging to the SOS regulon. Bacteria devoid of the SeqA protein, which preferentially binds hemimethylated DNA, were found to have a transcriptional profile almost identical to WT bacteria overexpressing Dam methyltransferase.

A nomenclature for restriction enzymes, DNA methyltransferases, homing endonucleases and their genes.

A nomenclature is described for restriction endonucleases, DNA methyltransferases, homing endonucleases and related genes and gene products. It provides explicit categories for the many different Type II enzymes now identified and provides a system for naming the putative genes found by sequence analysis of microbial genomes.

Nitric oxide-induced homologous recombination in Escherichia coli is promoted by DNA glycosylases.

Nitric oxide (NO*) is involved in neurotransmission, inflammation, and many other biological processes. Exposure of cells to NO* leads to DNA damage, including formation of deaminated and oxidized bases. Apurinic/apyrimidinic (AP) endonuclease-deficient cells are sensitive to NO* toxicity, which indicates that base excision repair (BER) intermediates are being generated.

MutS preferentially recognizes cisplatin- over oxaliplatin-modified DNA.

Loss of mismatch repair leads to tumor resistance by desensitizing cells to specific DNA-damaging agents, including the anticancer drug cisplatin. Cisplatin analogs with a diamminocyclohexane (DACH) carrier ligand, such as oxaliplatin and Pt(DACH)Cl(2), do not elicit resistance in mismatch repair-deficient cells and therefore present promising therapeutic agents. This study compared the interactions of the purified Escherichia coli mismatch repair protein MutS with DNA modified to contain cisplatin and DACH adducts.