Abstract
A genetic reversion assay to study C-to-T mutations within CG sites in DNA is described. It was used to demonstrate that the presence of HpaII methyltransferase (MTase) in Escherichia coli causes a substantial increase in C-to-T mutations at CG sites. This is similar to the known mutagenic effects of E. coli MTase Dcm within its own recognition sequence. With this genetic system, a homolog of an E. coli DNA repair gene in Haemophilus parainfluenzae was tested for antimutagenic activity. Unexpectedly, the homolog was found to have little effect on the reversion frequency. The system was also used to show that HpaII and SssI MTases can convert cytosine to uracil in vitro. These studies define 5-methylcytosine as an intrinsic mutagen and further elaborate the mutagenic potential of cytosine MTases.
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- Cooper D. N., Youssoufian H. The CpG dinucleotide and human genetic disease. Hum Genet. 1988 Feb;78(2):151–155. doi: 10.1007/BF00278187. [DOI] [PubMed] [Google Scholar]
- Coulondre C., Miller J. H., Farabaugh P. J., Gilbert W. Molecular basis of base substitution hotspots in Escherichia coli. Nature. 1978 Aug 24;274(5673):775–780. doi: 10.1038/274775a0. [DOI] [PubMed] [Google Scholar]
- Fishel R., Lescoe M. K., Rao M. R., Copeland N. G., Jenkins N. A., 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. doi: 10.1016/0092-8674(93)90546-3. [DOI] [PubMed] [Google Scholar]
- Giannelli F., Green P. M., High K. A., Lozier J. N., Lillicrap D. P., Ludwig M., Olek K., Reitsma P. H., Goossens M., Yoshioka A. Haemophilia B: database of point mutations and short additions and deletions. Nucleic Acids Res. 1990 Jul 25;18(14):4053–4059. doi: 10.1093/nar/18.14.4053. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hennecke F., Kolmar H., Bründl K., Fritz H. J. The vsr gene product of E. coli K-12 is a strand- and sequence-specific DNA mismatch endonuclease. Nature. 1991 Oct 24;353(6346):776–778. doi: 10.1038/353776a0. [DOI] [PubMed] [Google Scholar]
- Jones M., Wagner R., Radman M. Mismatch repair and recombination in E. coli. Cell. 1987 Aug 14;50(4):621–626. doi: 10.1016/0092-8674(87)90035-3. [DOI] [PubMed] [Google Scholar]
- Jones P. A., Rideout W. M., 3rd, Shen J. C., Spruck C. H., Tsai Y. C. Methylation, mutation and cancer. Bioessays. 1992 Jan;14(1):33–36. doi: 10.1002/bies.950140107. [DOI] [PubMed] [Google Scholar]
- Koeberl D. D., Bottema C. D., Ketterling R. P., Bridge P. J., Lillicrap D. P., Sommer S. S. Mutations causing hemophilia B: direct estimate of the underlying rates of spontaneous germ-line transitions, transversions, and deletions in a human gene. Am J Hum Genet. 1990 Aug;47(2):202–217. [PMC free article] [PubMed] [Google Scholar]
- Kulakauskas S., Barsomian J. M., Lubys A., Roberts R. J., Wilson G. G. Organization and sequence of the HpaII restriction-modification system and adjacent genes. Gene. 1994 May 3;142(1):9–15. doi: 10.1016/0378-1119(94)90348-4. [DOI] [PubMed] [Google Scholar]
- Kunkel T. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc Natl Acad Sci U S A. 1985 Jan;82(2):488–492. doi: 10.1073/pnas.82.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lieb M. Bacterial genes mutL, mutS, and dcm participate in repair of mismatches at 5-methylcytosine sites. J Bacteriol. 1987 Nov;169(11):5241–5246. doi: 10.1128/jb.169.11.5241-5246.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lieb M. Recombination in the lambda repressor gene: evidence that very short patch (VSP) mismatch correction restores a specific sequence. Mol Gen Genet. 1985;199(3):465–470. doi: 10.1007/BF00330759. [DOI] [PubMed] [Google Scholar]
- Lieb M. Specific mismatch correction in bacteriophage lambda crosses by very short patch repair. Mol Gen Genet. 1983;191(1):118–125. doi: 10.1007/BF00330898. [DOI] [PubMed] [Google Scholar]
- Lieb M. Spontaneous mutation at a 5-methylcytosine hotspot is prevented by very short patch (VSP) mismatch repair. Genetics. 1991 May;128(1):23–27. doi: 10.1093/genetics/128.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Marinus M. G., Morris N. R. Isolation of deoxyribonucleic acid methylase mutants of Escherichia coli K-12. J Bacteriol. 1973 Jun;114(3):1143–1150. doi: 10.1128/jb.114.3.1143-1150.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
- May M. S., Hattaman S. Deoxyribonucleic acid-cytosine methylation by host- and plasmid-controlled enzymes. J Bacteriol. 1975 Apr;122(1):129–138. doi: 10.1128/jb.122.1.129-138.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Prudhomme M., Méjean V., Martin B., Claverys J. P. Mismatch repair genes of Streptococcus pneumoniae: HexA confers a mutator phenotype in Escherichia coli by negative complementation. J Bacteriol. 1991 Nov;173(22):7196–7203. doi: 10.1128/jb.173.22.7196-7203.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Raleigh E. A., Wilson G. Escherichia coli K-12 restricts DNA containing 5-methylcytosine. Proc Natl Acad Sci U S A. 1986 Dec;83(23):9070–9074. doi: 10.1073/pnas.83.23.9070. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schlagman S., Hattman S., May M. S., Berger L. In vivo methylation by Escherichia coli K-12 mec+ deoxyribonucleic acid-cytosine methylase protects against in vitro cleavage by the RII restriction endonuclease (R. Eco RII). J Bacteriol. 1976 May;126(2):990–996. doi: 10.1128/jb.126.2.990-996.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Selker E. U. Premeiotic instability of repeated sequences in Neurospora crassa. Annu Rev Genet. 1990;24:579–613. doi: 10.1146/annurev.ge.24.120190.003051. [DOI] [PubMed] [Google Scholar]
- Shen J. C., Rideout W. M., 3rd, Jones P. A. High frequency mutagenesis by a DNA methyltransferase. Cell. 1992 Dec 24;71(7):1073–1080. doi: 10.1016/s0092-8674(05)80057-1. [DOI] [PubMed] [Google Scholar]
- Sohail A., Lieb M., Dar M., Bhagwat A. S. A gene required for very short patch repair in Escherichia coli is adjacent to the DNA cytosine methylase gene. J Bacteriol. 1990 Aug;172(8):4214–4221. doi: 10.1128/jb.172.8.4214-4221.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wyszynski M., Gabbara S., Bhagwat A. S. Cytosine deaminations catalyzed by DNA cytosine methyltransferases are unlikely to be the major cause of mutational hot spots at sites of cytosine methylation in Escherichia coli. Proc Natl Acad Sci U S A. 1994 Feb 15;91(4):1574–1578. doi: 10.1073/pnas.91.4.1574. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zhang X., Mathews C. K. Effect of DNA cytosine methylation upon deamination-induced mutagenesis in a natural target sequence in duplex DNA. J Biol Chem. 1994 Mar 11;269(10):7066–7069. [PubMed] [Google Scholar]