Role of damage-specific DNA polymerases in M13 phage mutagenesis induced by a major lipid peroxidation product trans-4-hydroxy-2-nonenal.

One of the major lipid peroxidation products trans-4-hydroxy-2-nonenal (HNE), forms cyclic propano- or ethenoadducts bearing six- or seven-carbon atom side chains to G>C≫A>T. To specify the role of SOS DNA polymerases in HNE-induced mutations, we tested survival and mutation spectra in the lacZα gene of M13mp18 phage, whose DNA was treated in vitro with HNE, and which was grown in uvrA(-)Escherichia coli strains, carrying one, two or all three SOS DNA polymerases. When Pol IV was the only DNA SOS polymerase in the bacterial host, survival of HNE-treated M13 DNA was similar to, but mutation frequency was lower than in the strain containing all SOS DNA polymerases.

Nucleotide excision repair and recombination are engaged in repair of trans-4-hydroxy-2-nonenal adducts to DNA bases in Escherichia coli.

One of the major products of lipid peroxidation is trans-4-hydroxy-2-nonenal (HNE). HNE forms highly mutagenic and genotoxic adducts to all DNA bases. Using M13 phage lacZ system, we studied the mutagenesis and repair of HNE treated phage DNA in E.

Mutagenic potency of MMS-induced 1meA/3meC lesions in E. coli.

The mutagenic activity of MMS in E. coli depends on the susceptibility of DNA bases to methylation and their repair by cellular defense systems. Among the lesions in methylated DNA is 1meA/3meC, which is recently recognized as being mutagenic.

Inducible SOS response system of DNA repair and mutagenesis in Escherichia coli.

Chromosomal DNA is exposed to continuous damage and repair. Cells contain a number of proteins and specific DNA repair systems that help maintain its correct structure. The SOS response was the first DNA repair system described in Escherichia coli induced upon treatment of bacteria with DNA damaging agents arrest DNA replication and cell division.

[Chemically methylated DNA repair in Escherichia coli--the role of alkB protein].

Methylating agents belong to mutagens occurring most frequently in our environment. They methylate mainly the nitrogen bases in DNA and RNA, affecting their functions. In E.

Mutator specificity of Escherichia coli alkB117 allele.

The Escherichia coli AlkB protein encoded by alkB gene was recently found to repair cytotoxic DNA lesions 1-methyladenine (1-meA) and 3-methylcytosine (3-meC) by using a novel iron-catalysed oxidative demethylation mechanism that protects the cell from the toxic effects of methylating agents. Mutation in alkB results in increased sensitivity to MMS and elevated level of MMS-induced mutations. The aim of this study was to analyse the mutational specificity of alkB117 in a system developed by J.

AlkB dioxygenase in preventing MMS-induced mutagenesis in Escherichia coli: effect of Pol V and AlkA proteins.

The deleterious effect of defective alkB allele encoding 1meA/3meC dioxygenase on reactivation of MMS-treated phage DNA has been frequently studied. Here, it is shown that: (i) AlkB protects the cells not only against the genotoxic but also against the potent mutagenic activity of MMS; (ii) mutations arising in alkB-defected strains are umuDC-dependent, and deletion of umuDC dramatically reduce MMS-induced mutations resulting from the presence of 1meA/3meC in DNA; (iii) specificity of MMS-induced argE3-->Arg+ reversions in AB1157 alkB-defective cells are predominantly AT-->TA transversions and GC-->AT transitions; (iv) overproduction of AlkA and the resultant decrease in 3meA residues in DNA dramatically reduce MMS-induced mutations. This reduction is most probably a secondary effect of AlkA due to a decrease in 3meA residues in DNA and, in consequence, suppression of SOS induction and Pol V expression.

Mutator activity and specificity of Escherichia coli dnaQ49 allele--effect of umuDC products.

The high fidelity of DNA replication in Escherichia coli is ensured by the alpha (DnaE) and epsilon (DnaQ) subunits of DNA polymerase providing insertion fidelity, 3'-->5' exonuclease proofreading activity, and by the dam-directed mismatch repair system. dnaQ49 is a recessive allele that confers a temperature-sensitive proofreading phenotype resulting in a high rate of spontaneous mutations and chronic induction of the SOS response. The aim of this study was to analyse the mutational specificity of dnaQ49 in umuDC and DeltaumuDC backgrounds at 28 and 37 degrees C in a system developed by J.

Effect of deletion of SOS-induced polymerases, pol II, IV, and V, on spontaneous mutagenesis in Escherichia coli mutD5.

The E. coli dnaQ gene encodes the epsilon subunit of DNA polymerase III (pol III) responsible for the proofreading activity of this polymerase. The mutD5 mutant of dnaQ chronically expresses the SOS response and exhibits a mutator phenotype.

E. coli BW535, a triple mutant for the DNA repair genes xth, nth, and nfo, chronically induces the SOS response.

A strong chronic induction of the SOS response system occurs in E. coli BW535, a strain defective in nth, nfo and xth genes, and hence severely deficient in the repair of abasic sites in DNA. This was shown here by visualization of filamentous growth of the BW535 strain and by measuring the level of beta-galactosidase expressed in BW535/pSK1002 in comparison to the AB1157/pSK1002 strain.

Lethality of visible light for Escherichia coli hemH1 mutants influence of defects in DNA repair.

The hemH gene encodes ferrochelatase, the final enzyme of the heme biosynthetic pathway. Defects of this enzyme lead to accumulation of protoporphyrin IX and an increase in reactive oxygen species, causing susceptibility to blue and white light in bacteria and protoporphyria in humans. Here we show that the photosensitivity of hemH1 strains is much increased when the bacteria are devoid of ability to repair abasic sites.

Induction of the SOS response in starved Escherichia coli.

The SOS system in Escherichia coli is induced in response to DNA damage and the arrest of DNA synthesis. Here we show that in AB1157 bacteria starved for arginine, conditions for induction of adaptive mutations, the LexA-dependent SOS system is induced, but that this occurs only when the bacteria resume growth and when the source of carbon is glycerol rather than glucose (glycerol, but not glucose, enables synthesis of cAMP). Therefore, we conclude that starved cells accumulate some lesions in DNA, which in growth conditions may trigger SOS induction by a process that is cAMP-dependent.