Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1996 Nov;64(11):4501–4507. doi: 10.1128/iai.64.11.4501-4507.1996

Effect of growth phase and acid shock on Helicobacter pylori cagA expression.

M Karita 1, M K Tummuru 1, H P Wirth 1, M J Blaser 1
PMCID: PMC174404  PMID: 8890198

Abstract

Helicobacter pylori strains possessing cagA are associated with peptic ulceration. To understand the regulation of expression of cagA, picB, associated with interleukin-8 induction, and ureA, encoding the small urease subunit, we created gene fusions of cagA, ureA, and picB of strain 3401, using a promoterless reporter (xylE). Expression of XylE after growth in broth culture revealed that basal levels of expression of cagA and urea in H. pylori were substantially greater than for picB. For cagA expression in stationary-phase cells, brief exposure to acid pH caused a significant increase in xylE expression compared with neutral pH. In contrast, expression of xylE in urea or picB decreased after parallel exposure to acid pH (pH 7 > 6 > 5 > 4), regardless of the growth phase. Expression of the CagA protein varied with growth phase and pH exposure in parallel with the observed transcriptional variation. The concentration of CagA in a cell membrane-enriched fraction after growth at pH 6 was significantly higher than after growth at pH 5 or 7. We conclude that the promoterless reporter xylE is useful for studying the regulation of gene expression in H. pylori and that regulation of CagA production occurs mainly at the transcriptional level.

Full Text

The Full Text of this article is available as a PDF (460.1 KB).

Selected References

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

  1. Andrew P. W., Roberts I. S. Construction of a bioluminescent mycobacterium and its use for assay of antimycobacterial agents. J Clin Microbiol. 1993 Sep;31(9):2251–2254. doi: 10.1128/jcm.31.9.2251-2254.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Barletta R. G., Kim D. D., Snapper S. B., Bloom B. R., Jacobs W. R., Jr Identification of expression signals of the mycobacteriophages Bxb1, L1 and TM4 using the Escherichia-Mycobacterium shuttle plasmids pYUB75 and pYUB76 designed to create translational fusions to the lacZ gene. J Gen Microbiol. 1992 Jan;138(1):23–30. doi: 10.1099/00221287-138-1-23. [DOI] [PubMed] [Google Scholar]
  4. Blaser M. J., Hopkins J. A., Berka R. M., Vasil M. L., Wang W. L. Identification and characterization of Campylobacter jejuni outer membrane proteins. Infect Immun. 1983 Oct;42(1):276–284. doi: 10.1128/iai.42.1.276-284.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Cave D. R., Vargas M. Effect of a Campylobacter pylori protein on acid secretion by parietal cells. Lancet. 1989 Jul 22;2(8656):187–189. doi: 10.1016/s0140-6736(89)90372-3. [DOI] [PubMed] [Google Scholar]
  7. Cooksey R. C., Crawford J. T., Jacobs W. R., Jr, Shinnick T. M. A rapid method for screening antimicrobial agents for activities against a strain of Mycobacterium tuberculosis expressing firefly luciferase. Antimicrob Agents Chemother. 1993 Jun;37(6):1348–1352. doi: 10.1128/aac.37.6.1348. [DOI] [PMC free article] [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. Cover T. L., Glupczynski Y., Lage A. P., Burette A., Tummuru M. K., Perez-Perez G. I., Blaser M. J. Serologic detection of infection with cagA+ Helicobacter pylori strains. J Clin Microbiol. 1995 Jun;33(6):1496–1500. doi: 10.1128/jcm.33.6.1496-1500.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cover T. L., Puryear W., Perez-Perez G. I., Blaser M. J. Effect of urease on HeLa cell vacuolation induced by Helicobacter pylori cytotoxin. Infect Immun. 1991 Apr;59(4):1264–1270. doi: 10.1128/iai.59.4.1264-1270.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Cover T. L., Tummuru M. K., Cao P., Thompson S. A., Blaser M. J. Divergence of genetic sequences for the vacuolating cytotoxin among Helicobacter pylori strains. J Biol Chem. 1994 Apr 8;269(14):10566–10573. [PubMed] [Google Scholar]
  13. Crabtree J. E., Taylor J. D., Wyatt J. I., Heatley R. V., Shallcross T. M., Tompkins D. S., Rathbone B. J. Mucosal IgA recognition of Helicobacter pylori 120 kDa protein, peptic ulceration, and gastric pathology. Lancet. 1991 Aug 10;338(8763):332–335. doi: 10.1016/0140-6736(91)90477-7. [DOI] [PubMed] [Google Scholar]
  14. Curcic R., Dhandayuthapani S., Deretic V. Gene expression in mycobacteria: transcriptional fusions based on xylE and analysis of the promoter region of the response regulator mtrA from Mycobacterium tuberculosis. Mol Microbiol. 1994 Sep;13(6):1057–1064. doi: 10.1111/j.1365-2958.1994.tb00496.x. [DOI] [PubMed] [Google Scholar]
  15. Das Gupta S. K., Bashyam M. D., Tyagi A. K. Cloning and assessment of mycobacterial promoters by using a plasmid shuttle vector. J Bacteriol. 1993 Aug;175(16):5186–5192. doi: 10.1128/jb.175.16.5186-5192.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Foster J. W., Bearson B. Acid-sensitive mutants of Salmonella typhimurium identified through a dinitrophenol lethal screening strategy. J Bacteriol. 1994 May;176(9):2596–2602. doi: 10.1128/jb.176.9.2596-2602.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Foster J. W. Salmonella acid shock proteins are required for the adaptive acid tolerance response. J Bacteriol. 1991 Nov;173(21):6896–6902. doi: 10.1128/jb.173.21.6896-6902.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  19. Jacobs W. R., Jr, Barletta R. G., Udani R., Chan J., Kalkut G., Sosne G., Kieser T., Sarkis G. J., Hatfull G. F., Bloom B. R. Rapid assessment of drug susceptibilities of Mycobacterium tuberculosis by means of luciferase reporter phages. Science. 1993 May 7;260(5109):819–822. doi: 10.1126/science.8484123. [DOI] [PubMed] [Google Scholar]
  20. Kangatharalingam N., Amy P. S. Helicobacter pylori comb. nov. Exhibits Facultative Acidophilism and Obligate Microaerophilism. Appl Environ Microbiol. 1994 Jun;60(6):2176–2179. doi: 10.1128/aem.60.6.2176-2179.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Karita M., Kouchiyama T., Okita K., Nakazawa T. New small animal model for human gastric Helicobacter pylori infection: success in both nude and euthymic mice. Am J Gastroenterol. 1991 Nov;86(11):1596–1603. [PubMed] [Google Scholar]
  22. Konyecsni W. M., Deretic V. Broad-host-range plasmid and M13 bacteriophage-derived vectors for promoter analysis in Escherichia coli and Pseudomonas aeruginosa. Gene. 1988 Dec 30;74(2):375–386. doi: 10.1016/0378-1119(88)90171-0. [DOI] [PubMed] [Google Scholar]
  23. Labigne A., Cussac V., Courcoux P. Shuttle cloning and nucleotide sequences of Helicobacter pylori genes responsible for urease activity. J Bacteriol. 1991 Mar;173(6):1920–1931. doi: 10.1128/jb.173.6.1920-1931.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mahan M. J., Slauch J. M., Mekalanos J. J. Selection of bacterial virulence genes that are specifically induced in host tissues. Science. 1993 Jan 29;259(5095):686–688. doi: 10.1126/science.8430319. [DOI] [PubMed] [Google Scholar]
  25. Mahan M. J., Tobias J. W., Slauch J. M., Hanna P. C., Collier R. J., Mekalanos J. J. Antibiotic-based selection for bacterial genes that are specifically induced during infection of a host. Proc Natl Acad Sci U S A. 1995 Jan 31;92(3):669–673. doi: 10.1073/pnas.92.3.669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Martin D. W., Schurr M. J., Mudd M. H., Govan J. R., Holloway B. W., Deretic V. Mechanism of conversion to mucoidy in Pseudomonas aeruginosa infecting cystic fibrosis patients. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8377–8381. doi: 10.1073/pnas.90.18.8377. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. McGowan C. C., Cover T. L., Blaser M. J. Helicobacter pylori and gastric acid: biological and therapeutic implications. Gastroenterology. 1996 Mar;110(3):926–938. doi: 10.1053/gast.1996.v110.pm8608904. [DOI] [PubMed] [Google Scholar]
  28. McGowan C. C., Cover T. L., Blaser M. J. The proton pump inhibitor omeprazole inhibits acid survival of Helicobacter pylori by a urease-independent mechanism. Gastroenterology. 1994 Sep;107(3):738–743. doi: 10.1016/0016-5085(94)90121-x. [DOI] [PubMed] [Google Scholar]
  29. Mohr C. D., Martin D. W., Konyecsni W. M., Govan J. R., Lory S., Deretic V. Role of the far-upstream sites of the algD promoter and the algR and rpoN genes in environmental modulation of mucoidy in Pseudomonas aeruginosa. J Bacteriol. 1990 Nov;172(11):6576–6580. doi: 10.1128/jb.172.11.6576-6580.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Murray A., Winter N., Lagranderie M., Hill D. F., Rauzier J., Timm J., Leclerc C., Moriarty K. M., Gheorghiu M., Gicquel B. Expression of Escherichia coli beta-galactosidase in Mycobacterium bovis BCG using an expression system isolated from Mycobacterium paratuberculosis which induced humoral and cellular immune responses. Mol Microbiol. 1992 Nov;6(22):3331–3342. doi: 10.1111/j.1365-2958.1992.tb02201.x. [DOI] [PubMed] [Google Scholar]
  31. Nakai C., Kagamiyama H., Nozaki M., Nakazawa T., Inouye S., Ebina Y., Nakazawa A. Complete nucleotide sequence of the metapyrocatechase gene on the TOI plasmid of Pseudomonas putida mt-2. J Biol Chem. 1983 Mar 10;258(5):2923–2928. [PubMed] [Google Scholar]
  32. Pérez-Pérez G. I., Olivares A. Z., Cover T. L., Blaser M. J. Characteristics of Helicobacter pylori variants selected for urease deficiency. Infect Immun. 1992 Sep;60(9):3658–3663. doi: 10.1128/iai.60.9.3658-3663.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Quigley E. M., Turnberg L. A. pH of the microclimate lining human gastric and duodenal mucosa in vivo. Studies in control subjects and in duodenal ulcer patients. Gastroenterology. 1987 Jun;92(6):1876–1884. doi: 10.1016/0016-5085(87)90619-6. [DOI] [PubMed] [Google Scholar]
  34. Schade C., Flemström G., Holm L. Hydrogen ion concentration in the mucus layer on top of acid-stimulated and -inhibited rat gastric mucosa. Gastroenterology. 1994 Jul;107(1):180–188. doi: 10.1016/0016-5085(94)90075-2. [DOI] [PubMed] [Google Scholar]
  35. Siegele D. A., Kolter R. Life after log. J Bacteriol. 1992 Jan;174(2):345–348. doi: 10.1128/jb.174.2.345-348.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Stover C. K., de la Cruz V. F., Fuerst T. R., Burlein J. E., Benson L. A., Bennett L. T., Bansal G. P., Young J. F., Lee M. H., Hatfull G. F. New use of BCG for recombinant vaccines. Nature. 1991 Jun 6;351(6326):456–460. doi: 10.1038/351456a0. [DOI] [PubMed] [Google Scholar]
  37. Taylor L. A., Rose R. E. A correction in the nucleotide sequence of the Tn903 kanamycin resistance determinant in pUC4K. Nucleic Acids Res. 1988 Jan 11;16(1):358–358. doi: 10.1093/nar/16.1.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. 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]
  39. Tummuru M. K., Sharma S. A., Blaser M. J. Helicobacter pylori picB, a homologue of the Bordetella pertussis toxin secretion protein, is required for induction of IL-8 in gastric epithelial cells. Mol Microbiol. 1995 Dec;18(5):867–876. doi: 10.1111/j.1365-2958.1995.18050867.x. [DOI] [PubMed] [Google Scholar]
  40. Wirth H. P., Wermuth B. Immunochemical characterization of aldo-keto reductases from human tissues. FEBS Lett. 1985 Aug 5;187(2):280–282. doi: 10.1016/0014-5793(85)81259-x. [DOI] [PubMed] [Google Scholar]
  41. Wirth H. P., Yang M., Karita M., Blaser M. J. Expression of the human cell surface glycoconjugates Lewis x and Lewis y by Helicobacter pylori isolates is related to cagA status. Infect Immun. 1996 Nov;64(11):4598–4605. doi: 10.1128/iai.64.11.4598-4605.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Yarchuk O., Jacques N., Guillerez J., Dreyfus M. Interdependence of translation, transcription and mRNA degradation in the lacZ gene. J Mol Biol. 1992 Aug 5;226(3):581–596. doi: 10.1016/0022-2836(92)90617-s. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES