Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1982 Sep;2(9):1096–1103. doi: 10.1128/mcb.2.9.1096

Mechanism of 2-aminopurine mutagenesis in mouse T-lymphosarcoma cells.

I W Caras, M A MacInnes, D H Persing, P Coffino, D W Martin Jr
PMCID: PMC369902  PMID: 6983647

Abstract

We investigated the mechanism of action of 2-aminopurine (Apur) in eucaryotic cells. By analogy with studies in procaryotic systems, the base analog is presumed to incorporate into DNA predominantly opposite T where, upon subsequent DNA replication, it can mispair with C, inducing an A:T leads to G:C transition. This model predicts that Apur-induced mutagenesis will be enhanced by factors that favor formation of Apur-C mispairs, e.g., high levels of dCTP or low levels of TTP. We describe the use of a mutant T-lymphosarcoma cell line, AraC-6-1, which has an abnormally high dCTP pool and a low TTP pool, to test this prediction. AraC-6-1 cells were three- to fivefold more mutable by Apur than their parental cell line, NSU-1. This enhanced mutability by Apur could not be explained by altered incorporation of 3H-labeled Apur, by generally impaired ability to repair DNA damage, or by a direct effect of Apur on the endogenous deoxynucleotide pools. The addition of 10 microM thymidine to the growth medium of AraC-6-1 cells lowered their high dCTP pool (two- to threefold), raised the TTP pool (two- to threefold), and abolished their enhanced mutability by Apur. Further manipulation to produce an abnormally high TTP/dCTP ratio suppressed Apur-induced mutagenesis (8- to 10-fold) in both AraC-6-1 and NSU-1 cells. These observations support the hypothesis that Apur induces A:T leads to G:C transitions in mammalian cells by a mispairing mechanism.

Full text

PDF
1096

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barrett J. C. Induction of gene mutation in and cell transformation of mammalian cells by modified purines: 2-aminopurine and 6-N-hydroxylaminopurine. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5685–5689. doi: 10.1073/pnas.78.9.5685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bradley M. O., Sharkey N. A. Mutagenicity of thymidine to cultured Chinese hamster cells. Nature. 1978 Aug 10;274(5671):607–608. doi: 10.1038/274607a0. [DOI] [PubMed] [Google Scholar]
  3. Chan T. S. Deoxyguanosine toxicity on lymphoid cells as a cause for immunosuppression in purine nucleoside phosphorylase deficiency. Cell. 1978 Jul;14(3):523–530. doi: 10.1016/0092-8674(78)90238-6. [DOI] [PubMed] [Google Scholar]
  4. Clive D., Johnson K. O., Spector J. F., Batson A. G., Brown M. M. Validation and characterization of the L5178Y/TK+/- mouse lymphoma mutagen assay system. Mutat Res. 1979 Jan;59(1):61–108. doi: 10.1016/0027-5107(79)90195-7. [DOI] [PubMed] [Google Scholar]
  5. Fersht A. R. Fidelity of replication of phage phi X174 DNA by DNA polymerase III holoenzyme: spontaneous mutation by misincorporation. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4946–4950. doi: 10.1073/pnas.76.10.4946. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Frederiksen S. Effect of 2-aminopurine and 2-aminopurine 2'-deoxyriboside on nucleic acid synthesis in Ehrlich ascites cells in vitro. Biochem Pharmacol. 1965 May;14(5):651–660. doi: 10.1016/0006-2952(65)90083-3. [DOI] [PubMed] [Google Scholar]
  7. Friedrich U., Coffino P. Mutagenesis in S49 mouse lymphoma cells: induction of resistance to ouabain, 6-thioguanine, and dibutyryl cyclic AMP. Proc Natl Acad Sci U S A. 1977 Feb;74(2):679–683. doi: 10.1073/pnas.74.2.679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Goodman M. F., Hopkins R., Gore W. C. 2-Aminopurine-induced mutagenesis in T4 bacteriophage: a model relating mutation frequency to 2-aminopurine incorporation in DNA. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4806–4810. doi: 10.1073/pnas.74.11.4806. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hibner U., Alberts B. M. Fidelity of DNA replication catalysed in vitro on a natural DNA template by the T4 bacteriophage multi-enzyme complex. Nature. 1980 May 29;285(5763):300–305. doi: 10.1038/285300a0. [DOI] [PubMed] [Google Scholar]
  10. Hopkins R. L., Goodman M. F. Deoxyribonucleotide pools, base pairing, and sequence configuration affecting bromodeoxyuridine- and 2-aminopurine-induced mutagenesis. Proc Natl Acad Sci U S A. 1980 Apr;77(4):1801–1805. doi: 10.1073/pnas.77.4.1801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hopkins R., Goodman M. F. Asymmetry in forming 2-aminopurine . hydroxymethylcytosine heteroduplexes; A model giving misincorporation frequencies and rounds of DNA replication from base-pair populations in vivo. J Mol Biol. 1979 Nov 25;135(1):1–22. doi: 10.1016/0022-2836(79)90337-1. [DOI] [PubMed] [Google Scholar]
  12. Horibata K., Harris A. W. Mouse myelomas and lymphomas in culture. Exp Cell Res. 1970 Apr;60(1):61–77. doi: 10.1016/0014-4827(70)90489-1. [DOI] [PubMed] [Google Scholar]
  13. Kunkel T. A., Loeb L. A. On the fidelity of DNA replication. Effect of divalent metal ion activators and deoxyrionucleoside triphosphate pools on in vitro mutagenesis. J Biol Chem. 1979 Jul 10;254(13):5718–5725. [PubMed] [Google Scholar]
  14. Kunkel T. A., Loeb L. A. On the fidelity of DNA replication. The accuracy of Escherichia coli DNA polymerase I in copying natural DNA in vitro. J Biol Chem. 1980 Oct 25;255(20):9961–9966. [PubMed] [Google Scholar]
  15. Meuth M., Green H. Induction of a deoxycytidineless state in cultured mammalian cells by bromodeoxyuridine. Cell. 1974 Jun;2(2):109–112. doi: 10.1016/0092-8674(74)90099-3. [DOI] [PubMed] [Google Scholar]
  16. Meuth M., L'Heureux-Huard N., Trudel M. Characterization of a mutator gene in Chinese hamster ovary cells. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6505–6509. doi: 10.1073/pnas.76.12.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Meuth M. Role of deoxynucleoside triphosphate pools in the cytotoxic and mutagenic effects of DNA alkylating agents. Somatic Cell Genet. 1981 Jan;7(1):89–102. doi: 10.1007/BF01544750. [DOI] [PubMed] [Google Scholar]
  18. Moore E. C., Hurlbert R. B. Regulation of mammalian deoxyribonucleotide biosynthesis by nucleotides as activators and inhibitors. J Biol Chem. 1966 Oct 25;241(20):4802–4809. [PubMed] [Google Scholar]
  19. Sibley C. H., Tomkins G. M. Isolation of lymphoma cell variants resistant to killing by glucocorticoids. Cell. 1974 Aug;2(4):213–220. doi: 10.1016/0092-8674(74)90013-0. [DOI] [PubMed] [Google Scholar]
  20. Ullman B., Gudas L. J., Clift S. M., Martin D. W., Jr Isolation and characterization of purine-nucleoside phosphorylase-deficient T-lymphoma cells and secondary mutants with altered ribonucleotide reductase: genetic model for immunodeficiency disease. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1074–1078. doi: 10.1073/pnas.76.3.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Ullman B., Gudas L. J., Cohen A., Martin D. W., Jr Deoxyadenosine metabolism and cytotoxicity in cultured mouse T lymphoma cells: a model for immunodeficiency disease. Cell. 1978 Jun;14(2):365–375. doi: 10.1016/0092-8674(78)90122-8. [DOI] [PubMed] [Google Scholar]
  22. Watanabe S. M., Goodman M. F. On the molecular basis of transition mutations: frequencies of forming 2-aminopurine.cytosine and adenine.cytosine base mispairs in vitro. Proc Natl Acad Sci U S A. 1981 May;78(5):2864–2868. doi: 10.1073/pnas.78.5.2864. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Weinberg G., Ullman B., Martin D. W., Jr Mutator phenotypes in mammalian cell mutants with distinct biochemical defects and abnormal deoxyribonucleoside triphosphate pools. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2447–2451. doi: 10.1073/pnas.78.4.2447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Weymouth L. A., Loeb L. A. Mutagenesis during in vitro DNA synthesis. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1924–1928. doi: 10.1073/pnas.75.4.1924. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES