Logo of jbacterPermissionsJournals.ASM.orgJournalJB ArticleJournal InfoAuthorsReviewers
J Bacteriol. 1994 Jun; 176(11): 3224–3230.
PMCID: PMC205492

DNA alkylation repair limits spontaneous base substitution mutations in Escherichia coli.


The Escherichia coli Ada and Ogt DNA methyltransferases (MTases) are known to transfer simple alkyl groups from O6-alkylguanine and O4-alkylthymine, directly restoring these alkylated DNA lesions to guanine and thymine. In addition to being exquisitely sensitive to the mutagenic effects of methylating agents, E. coli ada ogt null mutants display a higher spontaneous mutation rate than the wild type. Here, we determined which base substitution mutations are elevated in the MTase-deficient cells by monitoring the reversion of six mutated lacZ alleles that revert via each of the six possible base substitution mutations. During exponential growth, the spontaneous rate of G:C to A:T transitions and G:C to C:G transversions was elevated about fourfold in ada ogt double mutant versus wild-type E. coli. Furthermore, compared with the wild type, stationary populations of the MTase-deficient E. coli (under lactose selection) displayed increased G:C to A:T and A:T to G:C transitions (10- and 3-fold, respectively) and increased G:C to C:G, A:T to C:G, and A:T to T:A transversions (10-, 2.5-, and 1.7-fold, respectively). ada and ogt single mutants did not suffer elevated spontaneous mutation rates for any base substitution event, and the cloned ada and ogt genes each restored wild-type spontaneous mutation rates to the ada ogt MTase-deficient strains. We infer that both the Ada MTase and the Ogt MTase can repair the endogenously produced DNA lesions responsible for each of the five base substitution events that are elevated in MTase-deficient cells. Simple methylating and ethylating agents induced G:C to A:T and A:T to G:C transitions in these strains but did not significantly induce G:C to C:G, A:T to C:G, and A:T to T:A transversions. We deduce that S-adenosylmethionine (known to e a weak methylating agent) is not the only metabolite responsible for endogenous DNA alkylation and that at least some of the endogenous metabolites that cause O-alkyl DNA damage in E. coli are not simple methylating or ethylating agents.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.5M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Albertini AM, Hofer M, Calos MP, Miller JH. On the formation of spontaneous deletions: the importance of short sequence homologies in the generation of large deletions. Cell. 1982 Jun;29(2):319–328. [PubMed]
  • Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A. 1993 Sep 1;90(17):7915–7922. [PMC free article] [PubMed]
  • Aquilina G, Biondo R, Dogliotti E, Meuth M, Bignami M. Expression of the endogenous O6-methylguanine-DNA-methyltransferase protects Chinese hamster ovary cells from spontaneous G:C to A:T transitions. Cancer Res. 1992 Dec 1;52(23):6471–6475. [PubMed]
  • Barrows LR, Magee PN. Nonenzymatic methylation of DNA by S-adenosylmethionine in vitro. Carcinogenesis. 1982;3(3):349–351. [PubMed]
  • Basu AK, Essigmann JM. Site-specifically modified oligodeoxynucleotides as probes for the structural and biological effects of DNA-damaging agents. Chem Res Toxicol. 1988 Jan-Feb;1(1):1–18. [PubMed]
  • Beranek DT, Weis CC, Swenson DH. A comprehensive quantitative analysis of methylated and ethylated DNA using high pressure liquid chromatography. Carcinogenesis. 1980 Jul;1(7):595–606. [PubMed]
  • Cairns J, Foster PL. Adaptive reversion of a frameshift mutation in Escherichia coli. Genetics. 1991 Aug;128(4):695–701. [PMC free article] [PubMed]
  • Cairns J, Overbaugh J, Miller S. The origin of mutants. Nature. 1988 Sep 8;335(6186):142–145. [PubMed]
  • Calmels S, Ohshima H, Crespi M, Leclerc H, Cattoen C, Bartsch H. N-nitrosamine formation by microorganisms isolated from human gastric juice and urine: biochemical studies on bacteria-catalysed nitrosation. IARC Sci Publ. 1987;(84):391–395. [PubMed]
  • Chung CT, Niemela SL, Miller RH. One-step preparation of competent Escherichia coli: transformation and storage of bacterial cells in the same solution. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2172–2175. [PMC free article] [PubMed]
  • Coulondre C, Miller JH. Genetic studies of the lac repressor. III. Additional correlation of mutational sites with specific amino acid residues. J Mol Biol. 1977 Dec 15;117(3):525–567. [PubMed]
  • Coyle JT, Puttfarcken P. Oxidative stress, glutamate, and neurodegenerative disorders. Science. 1993 Oct 29;262(5134):689–695. [PubMed]
  • Cupples CG, Miller JH. A set of lacZ mutations in Escherichia coli that allow rapid detection of each of the six base substitutions. Proc Natl Acad Sci U S A. 1989 Jul;86(14):5345–5349. [PMC free article] [PubMed]
  • Diller L, Kassel J, Nelson CE, Gryka MA, Litwak G, Gebhardt M, Bressac B, Ozturk M, Baker SJ, Vogelstein B, et al. p53 functions as a cell cycle control protein in osteosarcomas. Mol Cell Biol. 1990 Nov;10(11):5772–5781. [PMC free article] [PubMed]
  • Drake JW. A constant rate of spontaneous mutation in DNA-based microbes. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):7160–7164. [PMC free article] [PubMed]
  • Echols H, Goodman MF. Fidelity mechanisms in DNA replication. Annu Rev Biochem. 1991;60:477–511. [PubMed]
  • Farr SB, D'Ari R, Touati D. Oxygen-dependent mutagenesis in Escherichia coli lacking superoxide dismutase. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8268–8272. [PMC free article] [PubMed]
  • Fearon ER, Vogelstein B. A genetic model for colorectal tumorigenesis. Cell. 1990 Jun 1;61(5):759–767. [PubMed]
  • Fishel R, Lescoe MK, Rao MR, Copeland NG, Jenkins NA, Garber J, Kane M, Kolodner R. The human mutator gene homolog MSH2 and its association with hereditary nonpolyposis colon cancer. Cell. 1993 Dec 3;75(5):1027–1038. [PubMed]
  • Foster PL. Directed mutation: between unicorns and goats. J Bacteriol. 1992 Mar;174(6):1711–1716. [PMC free article] [PubMed]
  • Foster PL. Adaptive mutation: the uses of adversity. Annu Rev Microbiol. 1993;47:467–504. [PMC free article] [PubMed]
  • Foster PL, Cairns J. Mechanisms of directed mutation. Genetics. 1992 Aug;131(4):783–789. [PMC free article] [PubMed]
  • Greenberg JT, Demple B. Overproduction of peroxide-scavenging enzymes in Escherichia coli suppresses spontaneous mutagenesis and sensitivity to redox-cycling agents in oxyR-mutants. EMBO J. 1988 Aug;7(8):2611–2617. [PMC free article] [PubMed]
  • Hall BG. Adaptive evolution that requires multiple spontaneous mutations. I. Mutations involving an insertion sequence. Genetics. 1988 Dec;120(4):887–897. [PMC free article] [PubMed]
  • Hall BG. Spectrum of mutations that occur under selective and non-selective conditions in E. coli. Genetica. 1991;84(2):73–76. [PubMed]
  • Horsfall MJ, Gordon AJ, Burns PA, Zielenska M, van der Vliet GM, Glickman BW. Mutational specificity of alkylating agents and the influence of DNA repair. Environ Mol Mutagen. 1990;15(2):107–122. [PubMed]
  • Lindahl T. Instability and decay of the primary structure of DNA. Nature. 1993 Apr 22;362(6422):709–715. [PubMed]
  • Lindahl T, Sedgwick B, Sekiguchi M, Nakabeppu Y. Regulation and expression of the adaptive response to alkylating agents. Annu Rev Biochem. 1988;57:133–157. [PubMed]
  • Loeb LA. Mutator phenotype may be required for multistage carcinogenesis. Cancer Res. 1991 Jun 15;51(12):3075–3079. [PubMed]
  • Loechler EL, Green CL, Essigmann JM. In vivo mutagenesis by O6-methylguanine built into a unique site in a viral genome. Proc Natl Acad Sci U S A. 1984 Oct;81(20):6271–6275. [PMC free article] [PubMed]
  • Luria SE, Delbrück M. Mutations of Bacteria from Virus Sensitivity to Virus Resistance. Genetics. 1943 Nov;28(6):491–511. [PMC free article] [PubMed]
  • Maki H, Sekiguchi M. MutT protein specifically hydrolyses a potent mutagenic substrate for DNA synthesis. Nature. 1992 Jan 16;355(6357):273–275. [PubMed]
  • Margison GP, Cooper DP, Potter PM. The E. coli ogt gene. Mutat Res. 1990 Nov-Dec;233(1-2):15–21. [PubMed]
  • Marnett LJ, Burcham PC. Endogenous DNA adducts: potential and paradox. Chem Res Toxicol. 1993 Nov-Dec;6(6):771–785. [PubMed]
  • Modrich P. Mechanisms and biological effects of mismatch repair. Annu Rev Genet. 1991;25:229–253. [PubMed]
  • Paik WK, Lee HW, Kim S. Non-enzymatic methylation of proteins with S-adenosyl-L-methionine. FEBS Lett. 1975 Oct 15;58(1):39–42. [PubMed]
  • Park JW, Ames BN. 7-Methylguanine adducts in DNA are normally present at high levels and increase on aging: analysis by HPLC with electrochemical detection. Proc Natl Acad Sci U S A. 1988 Oct;85(20):7467–7470. [PMC free article] [PubMed]
  • Parsons R, Li GM, Longley MJ, Fang WH, Papadopoulos N, Jen J, de la Chapelle A, Kinzler KW, Vogelstein B, Modrich P. Hypermutability and mismatch repair deficiency in RER+ tumor cells. Cell. 1993 Dec 17;75(6):1227–1236. [PubMed]
  • Patel DJ, Shapiro L, Kozlowski SA, Gaffney BL, Jones RA. Covalent carcinogenic O6-methylguanosine lesions in DNA. Structural studies of the O6 meG X A and O6meG X G interactions in dodecanucleotide duplexes. J Mol Biol. 1986 Apr 20;188(4):677–692. [PubMed]
  • Poltev VI, Steinberg SV. The role of structural water in the formation of nucleotide mispairs. J Biomol Struct Dyn. 1987 Oct;5(2):307–312. [PubMed]
  • Preston BD, Singer B, Loeb LA. Mutagenic potential of O4-methylthymine in vivo determined by an enzymatic approach to site-specific mutagenesis. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8501–8505. [PMC free article] [PubMed]
  • Rebeck GW, Samson L. Increased spontaneous mutation and alkylation sensitivity of Escherichia coli strains lacking the ogt O6-methylguanine DNA repair methyltransferase. J Bacteriol. 1991 Mar;173(6):2068–2076. [PMC free article] [PubMed]
  • Rydberg B, Lindahl T. Nonenzymatic methylation of DNA by the intracellular methyl group donor S-adenosyl-L-methionine is a potentially mutagenic reaction. EMBO J. 1982;1(2):211–216. [PMC free article] [PubMed]
  • Samson L. The suicidal DNA repair methyltransferases of microbes. Mol Microbiol. 1992 Apr;6(7):825–831. [PubMed]
  • Samson L, Thomale J, Rajewsky MF. Alternative pathways for the in vivo repair of O6-alkylguanine and O4-alkylthymine in Escherichia coli: the adaptive response and nucleotide excision repair. EMBO J. 1988 Jul;7(7):2261–2267. [PMC free article] [PubMed]
  • Sargentini NJ, Smith KC. Much of spontaneous mutagenesis in Escherichia coli is due to error-prone DNA repair: implications for spontaneous carcinogenesis. Carcinogenesis. 1981;2(9):863–872. [PubMed]
  • Shevell DE, Abou-Zamzam AM, Demple B, Walker GC. Construction of an Escherichia coli K-12 ada deletion by gene replacement in a recD strain reveals a second methyltransferase that repairs alkylated DNA. J Bacteriol. 1988 Jul;170(7):3294–3296. [PMC free article] [PubMed]
  • Shuker DE, Farmer PB. Relevance of urinary DNA adducts as markers of carcinogen exposure. Chem Res Toxicol. 1992 Jul-Aug;5(4):450–460. [PubMed]
  • Singer B, Dosanjh MK. Site-directed mutagenesis for quantitation of base-base interactions at defined sites. Mutat Res. 1990 Nov-Dec;233(1-2):45–51. [PubMed]
  • Smith TF, Sadler JR. The nature of lactose operator constitive mutations. J Mol Biol. 1971 Jul 28;59(2):273–305. [PubMed]
  • Solomon E, Borrow J, Goddard AD. Chromosome aberrations and cancer. Science. 1991 Nov 22;254(5035):1153–1160. [PubMed]
  • Storz G, Christman MF, Sies H, Ames BN. Spontaneous mutagenesis and oxidative damage to DNA in Salmonella typhimurium. Proc Natl Acad Sci U S A. 1987 Dec;84(24):8917–8921. [PMC free article] [PubMed]
  • Strauss BS. The origin of point mutations in human tumor cells. Cancer Res. 1992 Jan 15;52(2):249–253. [PubMed]
  • Tan BH, Bencsath FA, Gaubatz JW. Steady-state levels of 7-methylguanine increase in nuclear DNA of postmitotic mouse tissues during aging. Mutat Res. 1990 Sep-Nov;237(5-6):229–238. [PubMed]
  • Tsimis J, Yarosh DB. Adaptive response induction by bacterial catalysis of nitrosation. Environ Mol Mutagen. 1990;15(2):69–70. [PubMed]
  • Vaca CE, Wilhelm J, Harms-Ringdahl M. Interaction of lipid peroxidation products with DNA. A review. Mutat Res. 1988 Mar;195(2):137–149. [PubMed]
  • Vaughan P, Sedgwick B, Hall J, Gannon J, Lindahl T. Environmental mutagens that induce the adaptive response to alkylating agents in Escherichia coli. Carcinogenesis. 1991 Feb;12(2):263–268. [PubMed]
  • Xiao W, Samson L. In vivo evidence for endogenous DNA alkylation damage as a source of spontaneous mutation in eukaryotic cells. Proc Natl Acad Sci U S A. 1993 Mar 15;90(6):2117–2121. [PMC free article] [PubMed]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)


Save items

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • Gene
    Gene records that cite the current articles. Citations in Gene are added manually by NCBI or imported from outside public resources.
  • GEO Profiles
    GEO Profiles
    Gene Expression Omnibus (GEO) Profiles of molecular abundance data. The current articles are references on the Gene record associated with the GEO profile.
  • MedGen
    Related information in MedGen
  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem chemical substance records that cite the current articles. These references are taken from those provided on submitted PubChem chemical substance records.

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...