Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1997 Nov;179(21):6807–6815. doi: 10.1128/jb.179.21.6807-6815.1997

The Helicobacter pylori genome is modified at CATG by the product of hpyIM.

Q Xu 1, R M Peek Jr 1, G G Miller 1, M J Blaser 1
PMCID: PMC179612  PMID: 9352933

Abstract

To understand mechanisms of DNA methylation in Helicobacter pylori, a human pathogen associated with peptic ulcer disease and gastric adenocarcinoma, we cloned a putative DNA methyltransferase gene, hpyIM. This gene contains a 990-bp open reading frame encoding a 329-amino-acid protein, M.HpyI. Sequence analysis revealed that M.HpyI was closely related to CATG-recognizing adenine DNA methyltransferases, including M.NlaIII in N. lactamica. hpyIM was present in all H. pylori strains tested. DNA from wild-type H. pylori strains was resistant to digestion by SphI and NlaIII, which recognize DNA at sites containing CATG, whereas their isogenic hpyIM mutants were susceptible, indicating lack of modification. Overexpression of hpyIM in Escherichia coli rendered DNA from these cells resistant to NlaIII digestion, confirming the role of hpyIM in modifying CATG sites. We conclude that hpyIM encodes a DNA methyltransferase, M.HpyI, that is well conserved among diverse H. pylori strains and that modifies H. pylori genomes at CATG sites.

Full Text

The Full Text of this article is available as a PDF (917.5 KB).

Selected References

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

  1. Akopyanz N., Bukanov N. O., Westblom T. U., Berg D. E. PCR-based RFLP analysis of DNA sequence diversity in the gastric pathogen Helicobacter pylori. Nucleic Acids Res. 1992 Dec 11;20(23):6221–6225. doi: 10.1093/nar/20.23.6221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  3. Atherton J. C., Cao P., Peek R. M., Jr, Tummuru M. K., Blaser M. J., Cover T. L. Mosaicism in vacuolating cytotoxin alleles of Helicobacter pylori. Association of specific vacA types with cytotoxin production and peptic ulceration. J Biol Chem. 1995 Jul 28;270(30):17771–17777. doi: 10.1074/jbc.270.30.17771. [DOI] [PubMed] [Google Scholar]
  4. Barras F., Marinus M. G. The great GATC: DNA methylation in E. coli. Trends Genet. 1989 May;5(5):139–143. doi: 10.1016/0168-9525(89)90054-1. [DOI] [PubMed] [Google Scholar]
  5. Berg D. E., Logan R. P. Helicobacter pylori, individual host specificity and human disease. European Helicobacter Study Group Meeting, Copenhagen, October 16-19, 1996. Bioessays. 1997 Jan;19(1):86–90. doi: 10.1002/bies.950190114. [DOI] [PubMed] [Google Scholar]
  6. Blaser M. J., Parsonnet J. Parasitism by the "slow" bacterium Helicobacter pylori leads to altered gastric homeostasis and neoplasia. J Clin Invest. 1994 Jul;94(1):4–8. doi: 10.1172/JCI117336. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Braaten B. A., Nou X., Kaltenbach L. S., Low D. A. Methylation patterns in pap regulatory DNA control pyelonephritis-associated pili phase variation in E. coli. Cell. 1994 Feb 11;76(3):577–588. doi: 10.1016/0092-8674(94)90120-1. [DOI] [PubMed] [Google Scholar]
  8. Covacci A., Censini S., Bugnoli M., Petracca R., Burroni D., Macchia G., Massone A., Papini E., Xiang Z., Figura N. Molecular characterization of the 128-kDa immunodominant antigen of Helicobacter pylori associated with cytotoxicity and duodenal ulcer. Proc Natl Acad Sci U S A. 1993 Jun 15;90(12):5791–5795. doi: 10.1073/pnas.90.12.5791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cover T. L., Dooley C. P., Blaser M. J. Characterization of and human serologic response to proteins in Helicobacter pylori broth culture supernatants with vacuolizing cytotoxin activity. Infect Immun. 1990 Mar;58(3):603–610. doi: 10.1128/iai.58.3.603-610.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  12. Figura N., Guglielmetti P., Rossolini A., Barberi A., Cusi G., Musmanno R. A., Russi M., Quaranta S. Cytotoxin production by Campylobacter pylori strains isolated from patients with peptic ulcers and from patients with chronic gastritis only. J Clin Microbiol. 1989 Jan;27(1):225–226. doi: 10.1128/jcm.27.1.225-226.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gish W., States D. J. Identification of protein coding regions by database similarity search. Nat Genet. 1993 Mar;3(3):266–272. doi: 10.1038/ng0393-266. [DOI] [PubMed] [Google Scholar]
  14. Go M. F., Kapur V., Graham D. Y., Musser J. M. Population genetic analysis of Helicobacter pylori by multilocus enzyme electrophoresis: extensive allelic diversity and recombinational population structure. J Bacteriol. 1996 Jul;178(13):3934–3938. doi: 10.1128/jb.178.13.3934-3938.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Grilley M., Welsh K. M., Su S. S., Modrich P. Isolation and characterization of the Escherichia coli mutL gene product. J Biol Chem. 1989 Jan 15;264(2):1000–1004. [PubMed] [Google Scholar]
  16. Hill C., Miller L. A., Klaenhammer T. R. In vivo genetic exchange of a functional domain from a type II A methylase between lactococcal plasmid pTR2030 and a virulent bacteriophage. J Bacteriol. 1991 Jul;173(14):4363–4370. doi: 10.1128/jb.173.14.4363-4370.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hoke C., Vedros N. A. Characterization of atypical aerobic gram-negative cocci isolated from humans. J Clin Microbiol. 1982 May;15(5):906–914. doi: 10.1128/jcm.15.5.906-914.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Holland D., Wolk C. P. Identification and characterization of hetA, a gene that acts early in the process of morphological differentiation of heterocysts. J Bacteriol. 1990 Jun;172(6):3131–3137. doi: 10.1128/jb.172.6.3131-3137.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jiang Q., Hiratsuka K., Taylor D. E. Variability of gene order in different Helicobacter pylori strains contributes to genome diversity. Mol Microbiol. 1996 May;20(4):833–842. doi: 10.1111/j.1365-2958.1996.tb02521.x. [DOI] [PubMed] [Google Scholar]
  20. Kansau I., Raymond J., Bingen E., Courcoux P., Kalach N., Bergeret M., Braimi N., Dupont C., Labigne A. Genotyping of Helicobacter pylori isolates by sequencing of PCR products and comparison with the RAPD technique. Res Microbiol. 1996 Oct;147(8):661–669. doi: 10.1016/0923-2508(96)84023-x. [DOI] [PubMed] [Google Scholar]
  21. Karita M., Tummuru M. K., Wirth H. P., Blaser M. J. Effect of growth phase and acid shock on Helicobacter pylori cagA expression. Infect Immun. 1996 Nov;64(11):4501–4507. doi: 10.1128/iai.64.11.4501-4507.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kita K., Kotani H., Sugisaki H., Takanami M. The fokI restriction-modification system. I. Organization and nucleotide sequences of the restriction and modification genes. J Biol Chem. 1989 Apr 5;264(10):5751–5756. [PubMed] [Google Scholar]
  23. Kita K., Suisha M., Kotani H., Yanase H., Kato N. Cloning and sequence analysis of the StsI restriction-modification gene: presence of homology to FokI restriction-modification enzymes. Nucleic Acids Res. 1992 Aug 25;20(16):4167–4172. doi: 10.1093/nar/20.16.4167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Labbé D., Höltke H. J., Lau P. C. Cloning and characterization of two tandemly arranged DNA methyltransferase genes of Neisseria lactamica: an adenine-specific M.NlaIII and a cytosine-type methylase. Mol Gen Genet. 1990 Oct;224(1):101–110. doi: 10.1007/BF00259456. [DOI] [PubMed] [Google Scholar]
  25. Lauster R., Trautner T. A., Noyer-Weidner M. Cytosine-specific type II DNA methyltransferases. A conserved enzyme core with variable target-recognizing domains. J Mol Biol. 1989 Mar 20;206(2):305–312. doi: 10.1016/0022-2836(89)90480-4. [DOI] [PubMed] [Google Scholar]
  26. Lindahl T. DNA repair enzymes. Annu Rev Biochem. 1982;51:61–87. doi: 10.1146/annurev.bi.51.070182.000425. [DOI] [PubMed] [Google Scholar]
  27. Malone T., Blumenthal R. M., Cheng X. Structure-guided analysis reveals nine sequence motifs conserved among DNA amino-methyltransferases, and suggests a catalytic mechanism for these enzymes. J Mol Biol. 1995 Nov 3;253(4):618–632. doi: 10.1006/jmbi.1995.0577. [DOI] [PubMed] [Google Scholar]
  28. Meselson M., Yuan R. DNA restriction enzyme from E. coli. Nature. 1968 Mar 23;217(5134):1110–1114. doi: 10.1038/2171110a0. [DOI] [PubMed] [Google Scholar]
  29. Messer W., Noyer-Weidner M. Timing and targeting: the biological functions of Dam methylation in E. coli. Cell. 1988 Sep 9;54(6):735–737. doi: 10.1016/s0092-8674(88)90911-7. [DOI] [PubMed] [Google Scholar]
  30. Modrich P. Methyl-directed DNA mismatch correction. J Biol Chem. 1989 Apr 25;264(12):6597–6600. [PubMed] [Google Scholar]
  31. Morgan R. D., Camp R. R., Wilson G. G., Xu S. Y. Molecular cloning and expression of NlaIII restriction-modification system in E. coli. Gene. 1996 Dec 12;183(1-2):215–218. doi: 10.1016/s0378-1119(96)00561-6. [DOI] [PubMed] [Google Scholar]
  32. Nelson M., Raschke E., McClelland M. Effect of site-specific methylation on restriction endonucleases and DNA modification methyltransferases. Nucleic Acids Res. 1993 Jul 1;21(13):3139–3154. doi: 10.1093/nar/21.13.3139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Phadnis S. H., Westblom T. U., Normark S. Molecular cloning of Helicobacter pylori DNA: important differences between mcrBC deletion host strains. Mol Microbiol. 1993 Dec;10(5):1151–1151. doi: 10.1111/j.1365-2958.1993.tb00984.x. [DOI] [PubMed] [Google Scholar]
  34. Roberts D., Hoopes B. C., McClure W. R., Kleckner N. IS10 transposition is regulated by DNA adenine methylation. Cell. 1985 Nov;43(1):117–130. doi: 10.1016/0092-8674(85)90017-0. [DOI] [PubMed] [Google Scholar]
  35. Short J. M., Fernandez J. M., Sorge J. A., Huse W. D. Lambda ZAP: a bacteriophage lambda expression vector with in vivo excision properties. Nucleic Acids Res. 1988 Aug 11;16(15):7583–7600. doi: 10.1093/nar/16.15.7583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Smith D. B., Johnson K. S. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. doi: 10.1016/0378-1119(88)90005-4. [DOI] [PubMed] [Google Scholar]
  37. Smith D. W., Garland A. M., Herman G., Enns R. E., Baker T. A., Zyskind J. W. Importance of state of methylation of oriC GATC sites in initiation of DNA replication in Escherichia coli. EMBO J. 1985 May;4(5):1319–1326. doi: 10.1002/j.1460-2075.1985.tb03779.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Smith H. O., Annau T. M., Chandrasegaran S. Finding sequence motifs in groups of functionally related proteins. Proc Natl Acad Sci U S A. 1990 Jan;87(2):826–830. doi: 10.1073/pnas.87.2.826. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Staden R. Automation of the computer handling of gel reading data produced by the shotgun method of DNA sequencing. Nucleic Acids Res. 1982 Aug 11;10(15):4731–4751. doi: 10.1093/nar/10.15.4731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  41. Sugisaki H., Kita K., Takanami M. The FokI restriction-modification system. II. Presence of two domains in FokI methylase responsible for modification of different DNA strands. J Biol Chem. 1989 Apr 5;264(10):5757–5761. [PubMed] [Google Scholar]
  42. Taylor D. E., Eaton M., Chang N., Salama S. M. Construction of a Helicobacter pylori genome map and demonstration of diversity at the genome level. J Bacteriol. 1992 Nov;174(21):6800–6806. doi: 10.1128/jb.174.21.6800-6806.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tomb J. F., White O., Kerlavage A. R., Clayton R. A., Sutton G. G., Fleischmann R. D., Ketchum K. A., Klenk H. P., Gill S., Dougherty B. A. The complete genome sequence of the gastric pathogen Helicobacter pylori. Nature. 1997 Aug 7;388(6642):539–547. doi: 10.1038/41483. [DOI] [PubMed] [Google Scholar]
  44. Tummuru M. K., Cover T. L., Blaser M. J. Cloning and expression of a high-molecular-mass major antigen of Helicobacter pylori: evidence of linkage to cytotoxin production. Infect Immun. 1993 May;61(5):1799–1809. doi: 10.1128/iai.61.5.1799-1809.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wilson G. G., Murray N. E. Restriction and modification systems. Annu Rev Genet. 1991;25:585–627. doi: 10.1146/annurev.ge.25.120191.003101. [DOI] [PubMed] [Google Scholar]
  46. Woodcock D. M., Crowther P. J., Doherty J., Jefferson S., DeCruz E., Noyer-Weidner M., Smith S. S., Michael M. Z., Graham M. W. Quantitative evaluation of Escherichia coli host strains for tolerance to cytosine methylation in plasmid and phage recombinants. Nucleic Acids Res. 1989 May 11;17(9):3469–3478. doi: 10.1093/nar/17.9.3469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Zhang Y., Nelson M., Nietfeldt J. W., Burbank D. E., Van Etten J. L. Characterization of Chlorella virus PBCV-1 CviAII restriction and modification system. Nucleic Acids Res. 1992 Oct 25;20(20):5351–5356. doi: 10.1093/nar/20.20.5351. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES