Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1994 Mar;62(3):1046–1051. doi: 10.1128/iai.62.3.1046-1051.1994

Cloning, sequencing, and expression of the Escherichia coli cytolethal distending toxin genes.

C L Pickett 1, D L Cottle 1, E C Pesci 1, G Bikah 1
PMCID: PMC186222  PMID: 8112838

Abstract

A limited number of Escherichia coli isolates which produce an apparently novel toxin, termed cytolethal distending toxin (CDT), have been reported. The toxic activity produced by these strains causes certain cultured cell lines to become slowly distended and then disintegrate. DNA was isolated from the CDT-producing E. coli strain, 9142-88, and cloned into a cosmid vector. Plasmid DNA from a toxin-positive transductant was further subcloned until a plasmid with a 4-kb insert which still encoded the toxin activity was obtained. Nucleotide sequencing of a portion of this insert revealed the presence of three adjacent open reading frames. Further subcloning and deletion analysis suggested that the products of all three open reading frames may be required for toxin activity. Minicell experiments identified the products of all three open reading frames. The three proteins had predicted sizes of 27,753,29,531, and 19,938 Da, and all three appeared to have strong consensus leader sequences. None of the three predicted proteins had significant homology to known proteins.

Full text

PDF
1046

Images in this article

1049Image
on p.1049

Selected References

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

  1. Adler H. I., Fisher W. D., Cohen A., Hardigree A. A. MINIATURE escherichia coli CELLS DEFICIENT IN DNA. Proc Natl Acad Sci U S A. 1967 Feb;57(2):321–326. doi: 10.1073/pnas.57.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bouzari S., Varghese A. Cytolethal distending toxin (CLDT) production by enteropathogenic Escherichia coli (EPEC). FEMS Microbiol Lett. 1990 Sep 1;59(1-2):193–198. doi: 10.1016/0378-1097(90)90055-u. [DOI] [PubMed] [Google Scholar]
  3. Cohen S. N., Chang A. C., Hsu L. Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci U S A. 1972 Aug;69(8):2110–2114. doi: 10.1073/pnas.69.8.2110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Hohn B., Collins J. A small cosmid for efficient cloning of large DNA fragments. Gene. 1980 Nov;11(3-4):291–298. doi: 10.1016/0378-1119(80)90069-4. [DOI] [PubMed] [Google Scholar]
  5. Jobling M. G., Holmes R. K. Construction of vectors with the p15a replicon, kanamycin resistance, inducible lacZ alpha and pUC18 or pUC19 multiple cloning sites. Nucleic Acids Res. 1990 Sep 11;18(17):5315–5316. doi: 10.1093/nar/18.17.5315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Johnson W. M., Lior H. A new heat-labile cytolethal distending toxin (CLDT) produced by Campylobacter spp. Microb Pathog. 1988 Feb;4(2):115–126. doi: 10.1016/0882-4010(88)90053-8. [DOI] [PubMed] [Google Scholar]
  7. Johnson W. M., Lior H. A new heat-labile cytolethal distending toxin (CLDT) produced by Escherichia coli isolates from clinical material. Microb Pathog. 1988 Feb;4(2):103–113. doi: 10.1016/0882-4010(88)90052-6. [DOI] [PubMed] [Google Scholar]
  8. Lugtenberg B., Meijers J., Peters R., van der Hoek P., van Alphen L. Electrophoretic resolution of the "major outer membrane protein" of Escherichia coli K12 into four bands. FEBS Lett. 1975 Oct 15;58(1):254–258. doi: 10.1016/0014-5793(75)80272-9. [DOI] [PubMed] [Google Scholar]
  9. Meagher R. B., Tait R. C., Betlach M., Boyer H. W. Protein expression in E. coli minicells by recombinant plasmids. Cell. 1977 Mar;10(3):521–536. doi: 10.1016/0092-8674(77)90039-3. [DOI] [PubMed] [Google Scholar]
  10. Nagel de Zwaig R., Luria S. E. Genetics and physiology of colicin-tolerant mutants of Escherichia coli. J Bacteriol. 1967 Oct;94(4):1112–1123. doi: 10.1128/jb.94.4.1112-1123.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Pickett C. L., Twiddy E. M., Belisle B. W., Holmes R. K. Cloning of genes that encode a new heat-labile enterotoxin of Escherichia coli. J Bacteriol. 1986 Feb;165(2):348–352. doi: 10.1128/jb.165.2.348-352.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Pugsley A. P. The complete general secretory pathway in gram-negative bacteria. Microbiol Rev. 1993 Mar;57(1):50–108. doi: 10.1128/mr.57.1.50-108.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Scott D. A., Kaper J. B. Cloning and sequencing of the genes encoding Escherichia coli cytolethal distending toxin. Infect Immun. 1994 Jan;62(1):244–251. doi: 10.1128/iai.62.1.244-251.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Yamaguchi K., Yu F., Inouye M. A single amino acid determinant of the membrane localization of lipoproteins in E. coli. Cell. 1988 May 6;53(3):423–432. doi: 10.1016/0092-8674(88)90162-6. [DOI] [PubMed] [Google Scholar]

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

RESOURCES