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. 1989 Sep;123(1):123–129. doi: 10.1093/genetics/123.1.123

Sequence Analysis of N-Ethyl-N-Nitrosourea-Induced Vermilion Mutations in Drosophila Melanogaster

A Pastink 1, C Vreeken 1, MJM Nivard 1, L L Searles 1, E W Vogel 1
PMCID: PMC1203775  PMID: 2572507

Abstract

The mutational specificity of N-ethyl-N-nitrosourea (ENU) was determined in Drosophila melanogaster using the vermilion locus as a target gene. 25 mutants (16 F(1) and 9 F(2) mutants) were cloned and sequenced. Only base-pair changes were observed; three of the mutants represented double base substitutions. Transition mutations were the most prominent sequence change: 61% were GC->AT and 18% AT->GC substitutions. Both sequence changes can be explained by the miscoding properties of the modified guanine and thymine bases. A strong bias of neighboring bases on the occurrence of the GC->AT transitions or a strand preference of both types of transition mutations was not observed. The spectrum of ENU mutations in D. melanogaster includes a significant fraction (21%) of transversion mutations. Our data indicate that like in other prokaryotic and eukaryotic systems also in D. melanogaster the O(6)-ethylguanine adduct is the most prominent premutational lesion after ENU treatment. The strong contribution of the O(6)-ethylguanine adduct to the mutagenicity of ENU possibly explains the absence of distinct differences between the type of mutations observed in the F(1) and F(2) mutants. Although the latter arise later during development, the spectrum of mosaic mutations is also dominated by GC->AT transition mutations.

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Selected References

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  1. Baillie D. L., Chovnick A. Studies on the genetic control of tryptophan pyrrolase in Drosophila melanogaster. Mol Gen Genet. 1971;112(4):341–353. doi: 10.1007/BF00334435. [DOI] [PubMed] [Google Scholar]
  2. Batzer M. A., Tedeschi B., Fossett N. G., Tucker A., Kilroy G., Arbour P., Lee W. R. Spectra of molecular changes induced in DNA of Drosophila spermatozoa by 1-ethyl-1-nitrosourea and X-rays. Mutat Res. 1988 May;199(1):255–268. doi: 10.1016/0027-5107(88)90253-9. [DOI] [PubMed] [Google Scholar]
  3. Beranek D. T., Weis C. C., Swenson D. H. A comprehensive quantitative analysis of methylated and ethylated DNA using high pressure liquid chromatography. Carcinogenesis. 1980 Jul;1(7):595–606. doi: 10.1093/carcin/1.7.595. [DOI] [PubMed] [Google Scholar]
  4. Burns P. A., Gordon A. J., Glickman B. W. Influence of neighbouring base sequence on N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis in the lacI gene of Escherichia coli. J Mol Biol. 1987 Apr 5;194(3):385–390. doi: 10.1016/0022-2836(87)90668-1. [DOI] [PubMed] [Google Scholar]
  5. Burns P. A., Gordon A. J., Glickman B. W. Mutational specificity of N-methyl-N-nitrosourea in the lacI gene of Escherichia coli. Carcinogenesis. 1988 Sep;9(9):1607–1610. doi: 10.1093/carcin/9.9.1607. [DOI] [PubMed] [Google Scholar]
  6. Coté B., Bender W., Curtis D., Chovnick A. Molecular mapping of the rosy locus in Drosophila melanogaster. Genetics. 1986 Apr;112(4):769–783. doi: 10.1093/genetics/112.4.769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davis R. W., Thomas M., Cameron J., St John T. P., Scherer S., Padgett R. A. Rapid DNA isolations for enzymatic and hybridization analysis. Methods Enzymol. 1980;65(1):404–411. doi: 10.1016/s0076-6879(80)65051-4. [DOI] [PubMed] [Google Scholar]
  8. DuBridge R. B., Tang P., Hsia H. C., Leong P. M., Miller J. H., Calos M. P. Analysis of mutation in human cells by using an Epstein-Barr virus shuttle system. Mol Cell Biol. 1987 Jan;7(1):379–387. doi: 10.1128/mcb.7.1.379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Eckert K. A., Ingle C. A., Klinedinst D. K., Drinkwater N. R. Molecular analysis of mutations induced in human cells by N-ethyl-N-nitrosourea. Mol Carcinog. 1988;1(1):50–56. doi: 10.1002/mc.2940010111. [DOI] [PubMed] [Google Scholar]
  10. Lacy L. R., Eisenberg M. T., Osgood C. J. Molecular analysis of chemically-induced mutations at the RpII215 locus of Drosophila melanogaster. Mutat Res. 1986 Aug;162(1):47–54. doi: 10.1016/0027-5107(86)90070-9. [DOI] [PubMed] [Google Scholar]
  11. Lee C. S., Curtis D., McCarron M., Love C., Gray M., Bender W., Chovnick A. Mutations affecting expression of the rosy locus in Drosophila melanogaster. Genetics. 1987 May;116(1):55–66. doi: 10.1093/genetics/116.1.55. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lefevre G., Jr The eccentricity of vermilion deficiencies in Drosophila melanogaster. Genetics. 1969 Nov;63(3):589–600. doi: 10.1093/genetics/63.3.589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Levinson A., Silver D., Seed B. Minimal size plasmids containing an M13 origin for production of single-strand transducing particles. J Mol Appl Genet. 1984;2(6):507–517. [PubMed] [Google Scholar]
  14. Loeb L. A., Preston B. D. Mutagenesis by apurinic/apyrimidinic sites. Annu Rev Genet. 1986;20:201–230. doi: 10.1146/annurev.ge.20.120186.001221. [DOI] [PubMed] [Google Scholar]
  15. Loechler E. L., Green C. L., Essigmann J. M. 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. doi: 10.1073/pnas.81.20.6271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Marsh J. L., Erfle M., Wykes E. J. The pIC plasmid and phage vectors with versatile cloning sites for recombinant selection by insertional inactivation. Gene. 1984 Dec;32(3):481–485. doi: 10.1016/0378-1119(84)90022-2. [DOI] [PubMed] [Google Scholar]
  17. Nehls P., Rajewsky M. F. Ethylation of nucleophilic sites in DNA by N-ethyl-N-nitrosourea depends on chromatin structure and ionic strength. Mutat Res. 1985 Jun-Jul;150(1-2):13–21. doi: 10.1016/0027-5107(85)90096-x. [DOI] [PubMed] [Google Scholar]
  18. Pastink A., Vreeken C., Schalet A. P., Eeken J. C. DNA sequence analysis of X-ray-induced deletions at the white locus of Drosophila melanogaster. Mutat Res. 1988 Jan;207(1):23–28. doi: 10.1016/0165-7992(88)90006-1. [DOI] [PubMed] [Google Scholar]
  19. Pastink A., Vreeken C., Vogel E. W. The nature of N-ethyl-N-nitrosourea-induced mutations at the white locus of Drosophila melanogaster. Mutat Res. 1988 May;199(1):47–53. doi: 10.1016/0027-5107(88)90229-1. [DOI] [PubMed] [Google Scholar]
  20. Preston B. D., Singer B., Loeb L. A. 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. doi: 10.1073/pnas.83.22.8501. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Richardson K. K., Richardson F. C., Crosby R. M., Swenberg J. A., Skopek T. R. DNA base changes and alkylation following in vivo exposure of Escherichia coli to N-methyl-N-nitrosourea or N-ethyl-N-nitrosourea. Proc Natl Acad Sci U S A. 1987 Jan;84(2):344–348. doi: 10.1073/pnas.84.2.344. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Searles L. L., Voelker R. A. Molecular characterization of the Drosophila vermilion locus and its suppressible alleles. Proc Natl Acad Sci U S A. 1986 Jan;83(2):404–408. doi: 10.1073/pnas.83.2.404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Shen P., Huang H. V. Homologous recombination in Escherichia coli: dependence on substrate length and homology. Genetics. 1986 Mar;112(3):441–457. doi: 10.1093/genetics/112.3.441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wiseman R. W., Drinkwater N. R., Miller J. A., Miller E. C., Blomquist J. C. Apurinic/apyrimidinic site induction in supercoiled DNA and mutagenesis in Salmonella typhimurium TA100 by 1'-acetoxysafrole and related electrophilic alkenylbenzene derivatives. Carcinogenesis. 1986 Dec;7(12):2089–2093. doi: 10.1093/carcin/7.12.2089. [DOI] [PubMed] [Google Scholar]
  26. Zielenska M., Beranek D., Guttenplan J. B. Different mutational profiles induced by N-nitroso-N-ethylurea: effects of dose and error-prone DNA repair and correlations with DNA adducts. Environ Mol Mutagen. 1988;11(4):473–485. doi: 10.1002/em.2850110408. [DOI] [PubMed] [Google Scholar]

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