Abstract
Bordetella pertussis is the only bacteria] species which is known to produce pertussis toxin (PT); however, both Bordetella bronchiseptica and Bordetella parapertussis contain regions homologous to the ptx genes of B. pertussis that encode the toxin subunits. After finding that several children with B. parapertussis infections exhibited modest antibody titers to PT, we examined the ptx genes of both B. parapertussis and B. bronchiseptica to determine whether they would encode stable, functional proteins even though their promoters are thought to be inactive under the conditions that have been examined. We inserted a functional promoter directly upstream of the ptx-ptl region of both species and examined culture supernatants of the resulting strains for PT activity. Biologically active PT was found in the culture supernatants of both engineered species. The toxin encoded by the B. parapertussis ptx genes appeared more labile in culture supernatants than did toxin produced by either B. pertussis or the engineered strain of B. bronchiseptica. This lability might be due to the lack of a full-length S2 subunit. We also investigated the ptl genes of these species, which are necessary for the secretion of this toxin, and found that both B. bronchiseptica and B. parapertussis contain at least certain of these genes, including ptlE and ptlF. Moreover, B. bronchiseptica appeared to contain all essential ptl genes since the introduction of a functional promoter directly upstream of the ptx-ptl region resulted in both production and efficient secretion of toxin. These results indicate that despite a number of amino acid changes in the sequences of the toxins, the toxins encoded by B. bronchiseptica and B. parapertussis are active.
Full Text
The Full Text of this article is available as a PDF (267.1 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Aricò B., Gross R., Smida J., Rappuoli R. Evolutionary relationships in the genus Bordetella. Mol Microbiol. 1987 Nov;1(3):301–308. doi: 10.1111/j.1365-2958.1987.tb01936.x. [DOI] [PubMed] [Google Scholar]
- Aricò B., Rappuoli R. Bordetella parapertussis and Bordetella bronchiseptica contain transcriptionally silent pertussis toxin genes. J Bacteriol. 1987 Jun;169(6):2847–2853. doi: 10.1128/jb.169.6.2847-2853.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Barry E. M., Weiss A. A., Ehrmann I. E., Gray M. C., Hewlett E. L., Goodwin M. S. Bordetella pertussis adenylate cyclase toxin and hemolytic activities require a second gene, cyaC, for activation. J Bacteriol. 1991 Jan;173(2):720–726. doi: 10.1128/jb.173.2.720-726.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Burns D. L., Kenimer J. G., Manclark C. R. Role of the A subunit of pertussis toxin in alteration of Chinese hamster ovary cell morphology. Infect Immun. 1987 Jan;55(1):24–28. doi: 10.1128/iai.55.1.24-28.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Camilli A., Mekalanos J. J. Use of recombinase gene fusions to identify Vibrio cholerae genes induced during infection. Mol Microbiol. 1995 Nov;18(4):671–683. doi: 10.1111/j.1365-2958.1995.mmi_18040671.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cotter P. A., Miller J. F. BvgAS-mediated signal transduction: analysis of phase-locked regulatory mutants of Bordetella bronchiseptica in a rabbit model. Infect Immun. 1994 Aug;62(8):3381–3390. doi: 10.1128/iai.62.8.3381-3390.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- De Feyter R., Yang Y., Gabriel D. W. Gene-for-genes interactions between cotton R genes and Xanthomonas campestris pv. malvacearum avr genes. Mol Plant Microbe Interact. 1993 Mar-Apr;6(2):225–237. doi: 10.1094/mpmi-6-225. [DOI] [PubMed] [Google Scholar]
- Frank D. W., Parker C. D. Interaction of monoclonal antibodies with pertussis toxin and its subunits. Infect Immun. 1984 Oct;46(1):195–201. doi: 10.1128/iai.46.1.195-201.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gross R., Rappuoli R. Positive regulation of pertussis toxin expression. Proc Natl Acad Sci U S A. 1988 Jun;85(11):3913–3917. doi: 10.1073/pnas.85.11.3913. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Heininger U., Stehr K., Schmitt-Grohé S., Lorenz C., Rost R., Christenson P. D., Uberall M., Cherry J. D. Clinical characteristics of illness caused by Bordetella parapertussis compared with illness caused by Bordetella pertussis. Pediatr Infect Dis J. 1994 Apr;13(4):306–309. doi: 10.1097/00006454-199404000-00011. [DOI] [PubMed] [Google Scholar]
- Hewlett E. L., Sauer K. T., Myers G. A., Cowell J. L., Guerrant R. L. Induction of a novel morphological response in Chinese hamster ovary cells by pertussis toxin. Infect Immun. 1983 Jun;40(3):1198–1203. doi: 10.1128/iai.40.3.1198-1203.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Johnson F. D., Burns D. L. Detection and subcellular localization of three Ptl proteins involved in the secretion of pertussis toxin from Bordetella pertussis. J Bacteriol. 1994 Sep;176(17):5350–5356. doi: 10.1128/jb.176.17.5350-5356.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kenimer J. G., Kim K. J., Probst P. G., Manclark C. R., Burstyn D. G., Cowell J. L. Monoclonal antibodies to pertussis toxin: utilization as probes of toxin function. Hybridoma. 1989 Feb;8(1):37–51. doi: 10.1089/hyb.1989.8.37. [DOI] [PubMed] [Google Scholar]
- Kotob S. I., Hausman S. Z., Burns D. L. Localization of the promoter for the ptl genes of Bordetella pertussis, which encode proteins essential for secretion of pertussis toxin. Infect Immun. 1995 Aug;63(8):3227–3230. doi: 10.1128/iai.63.8.3227-3230.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Lee C. K., Roberts A., Perrin S. Expression of pertussis toxin in Bordetella bronchiseptica and Bordetella parapertussis carrying recombinant plasmids. Infect Immun. 1989 May;57(5):1413–1418. doi: 10.1128/iai.57.5.1413-1418.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Locht C., Geoffroy M. C., Renauld G. Common accessory genes for the Bordetella pertussis filamentous hemagglutinin and fimbriae share sequence similarities with the papC and papD gene families. EMBO J. 1992 Sep;11(9):3175–3183. doi: 10.1002/j.1460-2075.1992.tb05394.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Locht C., Keith J. M. Pertussis toxin gene: nucleotide sequence and genetic organization. Science. 1986 Jun 6;232(4755):1258–1264. doi: 10.1126/science.3704651. [DOI] [PubMed] [Google Scholar]
- Nicosia A., Perugini M., Franzini C., Casagli M. C., Borri M. G., Antoni G., Almoni M., Neri P., Ratti G., Rappuoli R. Cloning and sequencing of the pertussis toxin genes: operon structure and gene duplication. Proc Natl Acad Sci U S A. 1986 Jul;83(13):4631–4635. doi: 10.1073/pnas.83.13.4631. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nicosia A., Rappuoli R. Promoter of the pertussis toxin operon and production of pertussis toxin. J Bacteriol. 1987 Jun;169(6):2843–2846. doi: 10.1128/jb.169.6.2843-2846.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stein P. E., Boodhoo A., Armstrong G. D., Cockle S. A., Klein M. H., Read R. J. The crystal structure of pertussis toxin. Structure. 1994 Jan 15;2(1):45–57. doi: 10.1016/s0969-2126(00)00007-1. [DOI] [PubMed] [Google Scholar]
- Stein P. E., Boodhoo A., Armstrong G. D., Heerze L. D., Cockle S. A., Klein M. H., Read R. J. Structure of a pertussis toxin-sugar complex as a model for receptor binding. Nat Struct Biol. 1994 Sep;1(9):591–596. doi: 10.1038/nsb0994-591. [DOI] [PubMed] [Google Scholar]
- Stibitz S. Use of conditionally counterselectable suicide vectors for allelic exchange. Methods Enzymol. 1994;235:458–465. doi: 10.1016/0076-6879(94)35161-9. [DOI] [PubMed] [Google Scholar]
- Tamura M., Nogimori K., Murai S., Yajima M., Ito K., Katada T., Ui M., Ishii S. Subunit structure of islet-activating protein, pertussis toxin, in conformity with the A-B model. Biochemistry. 1982 Oct 26;21(22):5516–5522. doi: 10.1021/bi00265a021. [DOI] [PubMed] [Google Scholar]
- Vandamme P., Hommez J., Vancanneyt M., Monsieurs M., Hoste B., Cookson B., Wirsing von König C. H., Kersters K., Blackall P. J. Bordetella hinzii sp. nov., isolated from poultry and humans. Int J Syst Bacteriol. 1995 Jan;45(1):37–45. doi: 10.1099/00207713-45-1-37. [DOI] [PubMed] [Google Scholar]
- Walker M. J., Rohde M., Wehland J., Timmis K. N. Construction of minitransposons for constitutive and inducible expression of pertussis toxin in bvg-negative Bordetella bronchiseptica. Infect Immun. 1991 Nov;59(11):4238–4248. doi: 10.1128/iai.59.11.4238-4248.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weiss A. A., Hewlett E. L., Myers G. A., Falkow S. Pertussis toxin and extracytoplasmic adenylate cyclase as virulence factors of Bordetella pertussis. J Infect Dis. 1984 Aug;150(2):219–222. doi: 10.1093/infdis/150.2.219. [DOI] [PubMed] [Google Scholar]
- Weiss A. A., Johnson F. D., Burns D. L. Molecular characterization of an operon required for pertussis toxin secretion. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2970–2974. doi: 10.1073/pnas.90.7.2970. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weyant R. S., Hollis D. G., Weaver R. E., Amin M. F., Steigerwalt A. G., O'Connor S. P., Whitney A. M., Daneshvar M. I., Moss C. W., Brenner D. J. Bordetella holmesii sp. nov., a new gram-negative species associated with septicemia. J Clin Microbiol. 1995 Jan;33(1):1–7. doi: 10.1128/jcm.33.1.1-7.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]