Induction of mutations by replication of malondialdehyde modified M13 DNA in Escherichia coil: determination of the extent of DNA modification, genetic requirements for mutagenesis, and types of mutations induced

Abstract

The mutagenicity of the lipid peroxidation product, malondialdehyde (MDA), was measured in the lacZ alpha forward mutation assay using a recombinant M13 phage, M13MB102. Single-stranded M13MB102 DNA was reacted with MDA at neutral pH and the modified DNA was transformed into strains of Escherichia coli induced for the SOS response. Increasing concentrations of MDA led to an increase in lacZ alpha-mutations coincident with an increase in the level of the major MDA-deoxyguanosine adduct. Spontaneous and MDA-induced M13MB102 mutants were collected and the lacZ alpha target region was subjected to automated DNA sequence analysis. The most common sequence changes induced by MDA were base-pair substitutions (76%). Of these, 43% (29/68) were transversions, most of which were G-->T (24/29). Transitions account for 57% of the base-pair substitutions (39/68) and were comprised exclusively of C-->T (22/39) and A-->G (17/39). Frameshift mutations were identified in 16% of the induced mutants and were comprised of mainly single base additions occurring in runs of reiterated bases (11/14). The diversity of base-pair substitution and frameshift mutations induced by MDA at low levels of adduction suggests it may be an important contributor to endogenous mutagenesis and carcinogenesis in aerobic organisms.

Induction of frameshift and base pair substitution mutations by the major DNA adduct of the endogenous carcinogen malondialdehyde

“…The mutation spectra observed in our study reflect those reported for lacZ␣ mutations induced by treatment of M13 phage with MDA (20). In that study, 84% of mutations detected at G were base pair substitutions, with the remaining mutants comprising ϩG and ⌬CG mutations.…”

“…Z⌬M15]). LM102 and LM103 were constructed as described (20). The E. coli strain JM105 was used as the host bacterium for the replication of the M13MB102-1 genome on indicator plates.…”

MutS Recognition of Exocyclic DNA Adducts That Are Endogenous Products of Lipid Oxidation

“…MDA is mutagenic in bacterial and mammalian cell assays and carcinogenic in rodents (3)(4)(5)(6)(7). It also induces p53-independent cell cycle arrest at the G 1 /S and G 2 /M checkpoints (8).…”

Repair of Propanodeoxyguanosine by Nucleotide Excision Repair in Vivo and in Vitro

“…1) (2). These adducts have been detected at levels from 1500 -5000/ cell in liver DNA from healthy human beings (3,4) and appear to be highly mutagenic in site-specific mutagenesis assays (5)(6)(7)(8)(9)(10)(11).…”