Mechanisms of mutagenesis by exocyclic DNA adducts. Transfection of M13 viral DNA bearing a site-specific adduct shows that ethenocytosine is a highly efficient RecA-independent mutagenic noninstructional lesion

Abstract

It is widely accepted that mutagenic DNA lesions fall into two categories: mispairing lesions hydrogen bond with an incorrect incoming base, generally do not stop replication, and possess high mutagenic efficiency without any requirement for induced functions; noninstructional lesions lack accessible template information, act as strong blocks to DNA replication (and are therefore toxic), and their mutagenic effects are SOS-dependent. Our recent results show that ethenocytosine (epsilon C), a noninstructional exocyclic DNA lesion induced by vinyl chloride, may have unusual mutagenic properties. To obtain more definitive experimental evidence for the observed effects, we have introduced a single epsilon C residue at a specific site of coliphage M13AB28 replicative form DNA by a "single-stranded linker-ligation" technique. The resulting DNA was purified and transfected into appropriate recA+ or recA- Escherichia coli host cells. The effect of epsilon C on survival was determined from transfection efficiency. Both the frequency and specificity of mutations induced by epsilon C were determined by direct sequence analysis of randomly picked progeny phage plaques. The results indicated that epsilon C has little effect on the survival of M13 DNA. Approximately 30% of the progeny phage obtained by transfecting epsilon C DNA had a base substitution mutation precisely at the lesion site. No such mutations were observed in progeny plaques obtained by transfecting the control DNA construct. All epsilon C-induced mutations were either C-to-T transitions or C-to-A transversions. Neither survival nor mutagenic efficiency was significantly affected in recA- host cells.(ABSTRACT TRUNCATED AT 250 WORDS)

8-(Hydroxymethyl)-3,N4-etheno-C, a Potential Carcinogenic Glycidaldehyde Product, Miscodes In Vitro Using Mammalian DNA Polymerases

“…εC•T or εC•A were the main mispairs when E. coli polymerases were used (36)(37)(38). A later study using M13 viral DNA showed that 30% of progeny phage obtained by transfecting εC-DNA had a base substitution mutation at the lesion site (39). In agreement with the first reports, all these mutations were either C → T transitions or C → A transversions.…”

Functional recA, lexA, umuD, umuC, polA, and polB genes are not required for the Escherichia coli UVM response

“…Because of the etheno bridging of two of the three hydrogen-bonding positions, εC is expected to be a noninstructive lesion. The in vitro (56) and in vivo (27,28,49) template characteristics of εC are indeed those expected for such a lesion. For example, none of the four nucleotides are inserted by E. coli DNA pol I Klenow fragment opposite a site-specific εC residue borne on an oligodeoxyribonucleotide template under conditions in which guanine insertion opposite normal cytosine can be readily demonstrated on a control template (56).…”

“…Because of the etheno bridging of two of the three hydrogen-bonding positions, εC is expected to be a noninstructive lesion. The in vitro (56) and in vivo (27,28,49) template characteristics of εC …”

Damage of DNA and proteins by major lipid peroxidation products in genome stability

“…Ethenocytosine in site-directed mutagenesis experiments in E. coli and mammalian cells gave rise to 1.5 or 2% of targeted mutagenic events in non SOS-induced E. coli cells [55,58], 30% in SOS-induced ones, and 81% in simian kidney cells [58,59]. The predominant mutations generated by εC in both E. coli and COS cells were C:G → A:T transversions and C:G → T:A transitions.…”