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Molecular Genetics and Genomics

, Volume 267, Issue 1, pp 38–44 | Cite as

Organization and activation of the late promoters of φCTX, a cytotoxin-converting phage from Pseudomonas aeruginosa

  •  J. Alber
  •  F. Langewische
  •  A. Adebayo
  •  F. Lutz
Original Paper

Abstract.

The late genes of the temperate phage \( \phi {\rm CTX} \) of Pseudomonas aeruginosa are organized in an analogous fashion to the corresponding transcription units of the Escherichia coli P2 and P2-like phages. Sequence analysis of four putative late promoter regions, \( {\rm P}P_{\phi {\rm CTX}} \) , \({\rm P}O_{\phi {\rm CTX}} \) , \( {\rm P}V_{\phi {\rm CTX}} \) and \( {\rm P}F_{\phi {\rm CTX}} \) , reveals no similarity to σ70-type promoters or promoter consensus sequences found in Pseudomonas, indicating the apparent need for a phage-encoded protein to control the expression of \( \phi {\rm CTX} \) late genes. To elucidate the mode of expression of the late genes, we fused the putative late promoter regions to the promoterless lacZ α gene, which encodes the N-terminal part of β-galactosidase as a reporter enzyme, in the promoter-probe vector pME4510. The candidate transactivator gene orf34 was cloned into expression vector pHA10, to generate the plasmid pHA34. The two recombinant plasmids were introduced together into E. coli XL1-Blue and P. aeruginosa PAO1S-Lac. Our results demonstrate that in \( \phi {\rm CTX} \) three late promoters (\( {\rm P}P_{\phi {\rm CTX}} \) , \( {\rm P}O_{\phi {\rm CTX}} \) and \( {\rm P}F_{\phi {\rm CTX}} \) ) are activated upon induction by IPTG in PAO1S-Lac carrying the cloned promoters and pHA34. Deletions and base-pair substitutions obtained by PCR-mediated mutagenesis demonstrated that two conserved sequences, TTGTAG-N9-cTACAa and GcCGCGCGCGCGgC, are essential for effective late gene expression. Whereas the late promoters were active in P. aeruginosa, only weak β-galactosidase activity was obtained in E. coli.

Bacteriophage φCTX β-Galactosidase Late gene promoter Pseudomonas aeruginosa Transcriptional activation 

Copyright information

© Springer-Verlag 2002

Authors and Affiliations

  •  J. Alber
    • 1
  •  F. Langewische
    • 1
  •  A. Adebayo
    • 1
  •  F. Lutz
    • 1
  1. 1.Institut für Pharmakologie und Toxikologie, Justus-Liebig-Universität Giessen, Frankfurter Strasse 107, 35392 Giessen, Germany

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