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The role of mutation frequency decline and SOS repair systems in methyl methanesulfonate mutagenesis

100%

Grzesiuk E.

Acta Biochimica Polonica

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1998

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tom 45

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nr 2

Methyl methanesulfonate (MMS) is an SN2 type alkylating agent which predominantly methylates nitrogen atoms in purines. Among the methylated bases 3meA and 3meG are highly mutagenic and toxic. The excision of these lesions leads to the formation of apurinic (AP) sites and subsequently to AT-->TA or GC-->TA transversions. The in vivo method based on phenotypic analysis of Arg+ revertants of Escherichia coli K12 and sensitivity to T4 nonsense mutants has been used to estimate the specificity of MMS induced mutations. In the E. coli arg-his-thr- (AB1157) strain MMS induces argE3(oc)-->Arg+ revertants of which 70-80% arise by supL suppressor formation as a result of AT-->TA transversions. The remaining 20-30% arise by supB and supE(oc) suppressor formation as a result of GC-->AT transitions. The level of AT-->TA transversions decreases during starvation. This is a consequence of action of the repair mechanism called mutation frequency decline. This system which is a transcription coupled variant of nucleotide excision repair was discovered in UV induced mutations. We describe the mutation frequency decline phenomenon for MMS mutagenesis. MMS is a very efficient inducer of the SOS response and a umuDC dependent mutagen. In MMS treated E. coli cells mutated in umuDC genes the class of AT-->TA transversions dramatically diminishes. A plasmid bearing UmuD(D')C proteins can supplement chromosomal deletion of umuDC operon: a plasmid harbouring umuD'C is more efficient in comparison to that harbouring umuDC. Moreover, plasmids isolated from MMS treated and transiently starved E. coli AB1157 cells harbouring umuD(D')C genes have shown the repair of AP sites by a system which involves the UmuD'C or at least UmuD' protein.

2

Mechanisms of action of methyl methanesulfonate on Escherichia coli: mutagenesis. DNA damage and repair

100%

Janion C.

Postępy Biochemii

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1995

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tom 41

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nr 5

3

Mutator specificity of Escherichia coli alkB117 allele

100%

Nieminuszczy J., Janion C., Grzesiuk E.

Acta Biochimica Polonica

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2006

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tom 53

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nr 2

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.H. Miller involving two sets of E. coli lacZ mutants, CC101-106 allowing the identification of base pair substitutions, and CC107-CC111 indicating frameshift mutations. Of the six possible base substitutions, the presence of alkB117 allele led to an increased level of GC→AT transitions and GC→TA and AT→TA transversions. After MMS treatment the level of GC→AT transitions increased the most, 22-fold. Among frameshift mutations, the most numerous were -2CG, -1G, and -1A deletions and +1G insertion. MMS treatment appreciably increased all of the above types of frameshifts, with additional appearance of the +1A insertion.

4

Detection of alkylation damage in human lymphocyte DNA with the comet assay

80%

Collins A. R., Dusinska M., Horska A.

Acta Biochimica Polonica

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2001

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tom 48

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nr 3

The enzyme 3-methyladenine DNA glycosylase II (AlkA) is abacterial repair enzyme that acts preferentially at 3-methyladenine residues in DNA, releasing the damaged base. The resulting baseless sugars are alkali-labile, and under the conditions of the alkaline comet assay (single cell gel electrophoresis) they appear as DNA strand breaks. AlkA is not lesion-specific, but has a low activity even with undamaged bases. We have tested the enzyme at different concentrations to find conditions that maximise detection of alkylated bases with minimal attack on normal, undamaged DNA. AlkA detects damage in the DNA of cells treated with low concentrations of methyl methanesulphonate. We also find low background levels of alkylated bases in normal human lymphocytes.

Ograniczanie wyników

3 Acta Biochimica Polonica

1 Postępy Biochemii

2 Grzesiuk E.

2 Janion C.

1 Collins A. R.

1 Dusinska M.

1 Horska A.

1 Nieminuszczy J.

1 2006

1 2001

1 1998

1 1995

The role of mutation frequency decline and SOS repair systems in methyl methanesulfonate mutagenesis

Mechanisms of action of methyl methanesulfonate on Escherichia coli: mutagenesis. DNA damage and repair

Mutator specificity of Escherichia coli alkB117 allele

Detection of alkylation damage in human lymphocyte DNA with the comet assay