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. 1991 Oct 25;19(20):5525–5531. doi: 10.1093/nar/19.20.5525

Uracil-DNA glycosylase affects mismatch repair efficiency in transformation and bisulfite-induced mutagenesis in Streptococcus pneumoniae.

V Méjean 1, J C Devedjian 1, I Rives 1, G Alloing 1, J P Claverys 1
PMCID: PMC328952  PMID: 1945830

Abstract

The generalized mismatch repair system of Streptococcus pneumoniae (the Hex system) can eliminate base pair mismatches arising in heteroduplex DNA during transformation or by DNA polymerase errors during replication. Mismatch repair is most likely initiated at nicks or gaps. The present work was started to examine the hypothesis that strand discontinuities arising after removal of uracil by uracil DNA-glycosylase (Ung) can be utilised as strand discrimination signals. We show that mismatch repair efficiency is enhanced 3- to 6-fold when using uracil-containing DNA as donor in transformation. In order to assess the contribution of Ung to nascent strand discrimination for postreplication mismatch repair, we developed a positive selection procedure to isolate S. pneumoniae Ung- mutants. We succeeded in isolating Ung- mutants using this procedure based on chromosomal integration of uracil-containing hybrid DNA molecules. Cloning and characterization of the ung gene was achieved. Comparison of spontaneous mutation rates in strains either proficient or deficient in mismatch and/or uracil repair gave no support to the hypothesis that Ung plays a major role in targeting the Hex system to neosynthesized DNA strands. However Ung activity is responsible for the increased efficiency of mismatch repair observed in transformation with uracil-containing DNA. In addition Ung is involved in repair of bisulfite-treated transforming DNA.

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Selected References

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

  1. Alloing G., Trombe M. C., Claverys J. P. The ami locus of the gram-positive bacterium Streptococcus pneumoniae is similar to binding protein-dependent transport operons of gram-negative bacteria. Mol Microbiol. 1990 Apr;4(4):633–644. doi: 10.1111/j.1365-2958.1990.tb00632.x. [DOI] [PubMed] [Google Scholar]
  2. Aprelikova O., Jiricny J. Effect of uracil situated in the vicinity of a mispair on the directionality of mismatch correction in Escherichia coli. Nucleic Acids Res. 1991 Apr 11;19(7):1443–1447. doi: 10.1093/nar/19.7.1443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ballester S., Lopez P., Alonso J. C., Espinosa M., Lacks S. A. Selective advantage of deletions enhancing chloramphenicol acetyltransferase gene expression in Streptococcus pneumoniae plasmids. Gene. 1986;41(2-3):153–163. doi: 10.1016/0378-1119(86)90094-6. [DOI] [PubMed] [Google Scholar]
  4. Burgers P. M., Klein M. B. Selection by genetic transformation of a Saccharomyces cerevisiae mutant defective for the nuclear uracil-DNA-glycosylase. J Bacteriol. 1986 Jun;166(3):905–913. doi: 10.1128/jb.166.3.905-913.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chen J. D., Lacks S. A. Role of uracil-DNA glycosylase in mutation avoidance by Streptococcus pneumoniae. J Bacteriol. 1991 Jan;173(1):283–290. doi: 10.1128/jb.173.1.283-290.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Claverys J. P., Lacks S. A. Heteroduplex deoxyribonucleic acid base mismatch repair in bacteria. Microbiol Rev. 1986 Jun;50(2):133–165. doi: 10.1128/mr.50.2.133-165.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Claverys J. P., Méjean V. Strand targeting signal(s) for in vivo mutation avoidance by post-replication mismatch repair in Escherichia coli. Mol Gen Genet. 1988 Nov;214(3):574–578. doi: 10.1007/BF00330497. [DOI] [PubMed] [Google Scholar]
  8. Claverys J. P., Prats H., Vasseghi H., Gherardi M. Identification of Streptococcus pneumoniae mismatch repair genes by an additive transformation approach. Mol Gen Genet. 1984;196(1):91–96. doi: 10.1007/BF00334098. [DOI] [PubMed] [Google Scholar]
  9. Claverys J. P., Roger M., Sicard A. M. Excision and repair of mismatched base pairs in transformation of Streptococcus pneumoniae. Mol Gen Genet. 1980 Apr;178(1):191–201. doi: 10.1007/BF00267229. [DOI] [PubMed] [Google Scholar]
  10. Duncan B. K., Rockstroh P. A., Warner H. R. Escherichia coli K-12 mutants deficient in uracil-DNA glycosylase. J Bacteriol. 1978 Jun;134(3):1039–1045. doi: 10.1128/jb.134.3.1039-1045.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Duncan B. K., Weiss B. Specific mutator effects of ung (uracil-DNA glycosylase) mutations in Escherichia coli. J Bacteriol. 1982 Aug;151(2):750–755. doi: 10.1128/jb.151.2.750-755.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Frankel A. D., Duncan B. K., Hartman P. E. Nitrous acid damage to duplex deoxyribonucleic acid: distinction between deamination of cytosine residues and a novel mutational lesion. J Bacteriol. 1980 Apr;142(1):335–338. doi: 10.1128/jb.142.1.335-338.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gasc A. M., Sicard A. M., Claverys J. P. Repair of single- and multiple-substitution mismatches during recombination in Streptococcus pneumoniae. Genetics. 1989 Jan;121(1):29–36. doi: 10.1093/genetics/121.1.29. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Glickman B. W., Radman M. Escherichia coli mutator mutants deficient in methylation-instructed DNA mismatch correction. Proc Natl Acad Sci U S A. 1980 Feb;77(2):1063–1067. doi: 10.1073/pnas.77.2.1063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Grilley M., Holmes J., Yashar B., Modrich P. Mechanisms of DNA-mismatch correction. Mutat Res. 1990 Sep-Nov;236(2-3):253–267. doi: 10.1016/0921-8777(90)90009-t. [DOI] [PubMed] [Google Scholar]
  16. Guild W. R., Shoemaker N. B. Mismatch correction in pneumococcal transformation: donor length and hex-dependent marker efficiency. J Bacteriol. 1976 Jan;125(1):125–135. doi: 10.1128/jb.125.1.125-135.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Herman G. E., Modrich P. Escherichia coli K-12 clones that overproduce dam methylase are hypermutable. J Bacteriol. 1981 Jan;145(1):644–646. doi: 10.1128/jb.145.1.644-646.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Holmes J., Jr, Clark S., Modrich P. Strand-specific mismatch correction in nuclear extracts of human and Drosophila melanogaster cell lines. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5837–5841. doi: 10.1073/pnas.87.15.5837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kramer B., Kramer W., Williamson M. S., Fogel S. Heteroduplex DNA correction in Saccharomyces cerevisiae is mismatch specific and requires functional PMS genes. Mol Cell Biol. 1989 Oct;9(10):4432–4440. doi: 10.1128/mcb.9.10.4432. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kramer W., Kramer B., Williamson M. S., Fogel S. Cloning and nucleotide sequence of DNA mismatch repair gene PMS1 from Saccharomyces cerevisiae: homology of PMS1 to procaryotic MutL and HexB. J Bacteriol. 1989 Oct;171(10):5339–5346. doi: 10.1128/jb.171.10.5339-5346.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  22. Lacks S. A., Dunn J. J., Greenberg B. Identification of base mismatches recognized by the heteroduplex-DNA-repair system of Streptococcus pneumoniae. Cell. 1982 Dec;31(2 Pt 1):327–336. doi: 10.1016/0092-8674(82)90126-x. [DOI] [PubMed] [Google Scholar]
  23. Lacks S., Greenberg B. Deoxyribonucleases of Pneumococcus. J Biol Chem. 1967 Jul 10;242(13):3108–3120. [PubMed] [Google Scholar]
  24. Lindahl T. An N-glycosidase from Escherichia coli that releases free uracil from DNA containing deaminated cytosine residues. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3649–3653. doi: 10.1073/pnas.71.9.3649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lindahl T. Repair of intrinsic DNA lesions. Mutat Res. 1990 May;238(3):305–311. doi: 10.1016/0165-1110(90)90022-4. [DOI] [PubMed] [Google Scholar]
  26. Lyons S. M., Schendel P. F. Kinetics of methylation in Escherichia coli K-12. J Bacteriol. 1984 Jul;159(1):421–423. doi: 10.1128/jb.159.1.421-423.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Längle-Rouault F., Maenhaut-Michel G., Radman M. GATC sequences, DNA nicks and the MutH function in Escherichia coli mismatch repair. EMBO J. 1987 Apr;6(4):1121–1127. doi: 10.1002/j.1460-2075.1987.tb04867.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Marinus M. G. Location of DNA methylation genes on the Escherichia coli K-12 genetic map. Mol Gen Genet. 1973 Dec 14;127(1):47–55. doi: 10.1007/BF00267782. [DOI] [PubMed] [Google Scholar]
  29. Marinus M. G., Morris N. R. Biological function for 6-methyladenine residues in the DNA of Escherichia coli K12. J Mol Biol. 1974 May 15;85(2):309–322. doi: 10.1016/0022-2836(74)90366-0. [DOI] [PubMed] [Google Scholar]
  30. Martin B., Prats H., Claverys J. P. Cloning of the hexA mismatch-repair gene of Streptococcus pneumoniae and identification of the product. Gene. 1985;34(2-3):293–303. doi: 10.1016/0378-1119(85)90138-6. [DOI] [PubMed] [Google Scholar]
  31. Modrich P. Methyl-directed DNA mismatch correction. J Biol Chem. 1989 Apr 25;264(12):6597–6600. [PubMed] [Google Scholar]
  32. Morrison D. A., Trombe M. C., Hayden M. K., Waszak G. A., Chen J. D. Isolation of transformation-deficient Streptococcus pneumoniae mutants defective in control of competence, using insertion-duplication mutagenesis with the erythromycin resistance determinant of pAM beta 1. J Bacteriol. 1984 Sep;159(3):870–876. doi: 10.1128/jb.159.3.870-876.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Méjean V., Claverys J. P. Effect of mismatched base pairs on the fate of donor DNA in transformation of Streptococcus pneumoniae. Mol Gen Genet. 1984;197(3):467–471. doi: 10.1007/BF00329944. [DOI] [PubMed] [Google Scholar]
  34. Méjean V., Claverys J. P. Polarity of DNA entry in transformation of Streptococcus pneumoniae. Mol Gen Genet. 1988 Aug;213(2-3):444–448. doi: 10.1007/BF00339614. [DOI] [PubMed] [Google Scholar]
  35. Méjean V., Claverys J. P., Vasseghi H., Sicard A. M. Rapid cloning of specific DNA fragments of Streptococcus pneumoniae by vector integration into the chromosome followed by endonucleolytic excision. Gene. 1981 Nov;15(2-3):289–293. doi: 10.1016/0378-1119(81)90139-6. [DOI] [PubMed] [Google Scholar]
  36. Méjean V., Rives I., Claverys J. P. Nucleotide sequence of the Streptococcus pneumoniae ung gene encoding uracil-DNA glycosylase. Nucleic Acids Res. 1990 Nov 25;18(22):6693–6693. doi: 10.1093/nar/18.22.6693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Oeda K., Shimizu K., Sekiguchi M. An enzyme activity specific for nitrous acid-treated DNA in Escherichia coli. J Biochem. 1978 Nov;84(5):1165–1169. doi: 10.1093/oxfordjournals.jbchem.a132232. [DOI] [PubMed] [Google Scholar]
  38. Prats H., Martin B., Claverys J. P. The hexB mismatch repair gene of Streptococcus pneumoniae: characterisation, cloning and identification of the product. Mol Gen Genet. 1985;200(3):482–489. doi: 10.1007/BF00425735. [DOI] [PubMed] [Google Scholar]
  39. Prats H., Martin B., Pognonec P., Burger A. C., Claverys J. P. A plasmid vector allowing positive selection of recombinant plasmids in Streptococcus pneumoniae. Gene. 1985;39(1):41–48. doi: 10.1016/0378-1119(85)90105-2. [DOI] [PubMed] [Google Scholar]
  40. Prudhomme M., Martin B., Mejean V., Claverys J. P. Nucleotide sequence of the Streptococcus pneumoniae hexB mismatch repair gene: homology of HexB to MutL of Salmonella typhimurium and to PMS1 of Saccharomyces cerevisiae. J Bacteriol. 1989 Oct;171(10):5332–5338. doi: 10.1128/jb.171.10.5332-5338.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Puyet A., Greenberg B., Lacks S. A. The exoA gene of Streptococcus pneumoniae and its product, a DNA exonuclease with apurinic endonuclease activity. J Bacteriol. 1989 May;171(5):2278–2286. doi: 10.1128/jb.171.5.2278-2286.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Su S. S., Grilley M., Thresher R., Griffith J., Modrich P. Gap formation is associated with methyl-directed mismatch correction under conditions of restricted DNA synthesis. Genome. 1989;31(1):104–111. doi: 10.1139/g89-020. [DOI] [PubMed] [Google Scholar]
  43. Tiraby G., Sicard M. A. Integration efficiencies of spontaneous mutant alleles of amiA locus in pneumococcal transformation. J Bacteriol. 1973 Dec;116(3):1130–1135. doi: 10.1128/jb.116.3.1130-1135.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Tiraby J. G., Fox M. S. Marker discrimination and mutagen-induced alterations in pneumococcal transformation. Genetics. 1974 Jul;77(3):449–458. doi: 10.1093/genetics/77.3.449. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tiraby J. G., Fox M. S. Marker discrimination in transformation and mutation of pneumococcus. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3541–3545. doi: 10.1073/pnas.70.12.3541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Tye B. K., Chien J., Lehman I. R., Duncan B. K., Warner H. R. Uracil incorporation: a source of pulse-labeled DNA fragments in the replication of the Escherichia coli chromosome. Proc Natl Acad Sci U S A. 1978 Jan;75(1):233–237. doi: 10.1073/pnas.75.1.233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Vasseghi H., Claverys J. P. Amplification of a chimeric plasmid carrying an erythromycin-resistance determinant introduced into the genome of Streptococcus pneumoniae. Gene. 1983 Mar;21(3):285–292. doi: 10.1016/0378-1119(83)90012-4. [DOI] [PubMed] [Google Scholar]
  48. Wagner R., Jr, Meselson M. Repair tracts in mismatched DNA heteroduplexes. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4135–4139. doi: 10.1073/pnas.73.11.4135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Warner H. R., Duncan B. K., Garrett C., Neuhard J. Synthesis and metabolism of uracil-containing deoxyribonucleic acid in Escherichia coli. J Bacteriol. 1981 Feb;145(2):687–695. doi: 10.1128/jb.145.2.687-695.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Williamson M. S., Game J. C., Fogel S. Meiotic gene conversion mutants in Saccharomyces cerevisiae. I. Isolation and characterization of pms1-1 and pms1-2. Genetics. 1985 Aug;110(4):609–646. doi: 10.1093/genetics/110.4.609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Zimmermann F. Genetic effects of nitrous acid. Mutat Res. 1977;39(2):127–148. doi: 10.1016/0165-1110(77)90019-7. [DOI] [PubMed] [Google Scholar]

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