Skip to main content
SpringerLink
Log in

Site-specific integration of genes into hot spots for recombination flanking aadA in Tn21 transposons

  • Published:
Molecular and General Genetics MGG Aims and scope Submit manuscript

Summary

Tn21-related transposons are widespread among bacteria and carry various resistance determinants at preferential sites, hs1 and hs2. In an in vivo integrative recombination assay it was demonstrated that these hot spots direct the integration of aminoglycoside resistance genes like aadB from Klebsiella pneumoniae and aacAI from Serratia marcescens, in a recA − background. The maximum required recognition sequence which must be present in both the donor and recipient plasmids is 5′ CTAAAACAAAGTTA 3′ (hs2). The double-site-specific recombination occurred with a frequency of 10−5–10−6. The resulting structures include not only replicon fusion products but also more complex structures carrying two copies of the donor plasmid or simply the donor gene flanked by hs elements. hs1 and hs2 are thought to act as recognition sites for a trans-acting site-specific recombinase. By the use of Tn21 deletion derivatives, it has been shown that the recombinase is not encoded by Tn21. This new integrative recombination system is involved in the acquisition of new genes by Tn21-related transposons and their spread among bacterial populations.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Argos P, Landy A, Abremsky K, Egan JB, Haggard-Ljungquist E, Hoess RH, Kahn ML, Kalionis B, Narayana SVL, Pierson III LS, Sternberg N, Leong JM (1986) The integrase family of site-specific recombinases: Regional similarities and global diversity. EMBO J 5:433–440

    Google Scholar 

  • Avila P, Grinsted J, de la Cruz F (1988) Analysis of the variable endpoints generated by one-ended transposition of Tn21. J Bacteriol 170:1350–1353

    Google Scholar 

  • Brown NL, Misraq TK, Winnie JN Schmidt A, Seiff M, Silver S (1986) The nucleotide sequence of the mercuric resistance operons of plasmid R100 and transposon Tn501: further evidence for mer genes which enhance the activity of the mercuric ion detoxification systems. Mol Gen Genet 202:143–151

    Google Scholar 

  • Chater KF, Henderson DJ, Bibb MJ, Hopwood DA (1988) Genome fluxe in Streptomyces coelicolor and other streptomycetes and its possible relevance to the evolution of mobile antibiotic resistance determinants. Symp Soc Gen Microbiol 43:7–42

    Google Scholar 

  • Cohen SN, Chung AC, Hsu L (1972) Nonchromosomal antibiotic resistance in bacteria: Genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci USA 69:2110–2114

    CAS  PubMed  Google Scholar 

  • de la Cruz F, Grinsted J (1982) Genetic and molecular characterization of Tn21, a multiple resistance transposon from R100.1. J Bacteriol 151:222–228

    Google Scholar 

  • Dorman CJ, Higgins CF (1987) Fimbrial phase variation in Escherichia coli: Dependence on integration host factor and homologies with other site-specific recombinases. J Bacteriol 169:3840–3843

    Google Scholar 

  • Eisenstein I, Sweet DS, Vaughn V, Friedmann DI (1987) Integration host factor is required for the DNA inversion that controls phase variation in Escherichia coli. Proc Natl Acad Sci USA 84:6506–6510

    Google Scholar 

  • Grindley NDF, Reed RR (1985) Transpositional recombination in procaryotes. Annu Rev Biochem 54:863–896

    Google Scholar 

  • Guerry P, van Embden JD, Falkow S (1974) Molecular nature of two non-conjugative plasmids carrying drug resistance genes. J Bacteriol 117:619–630

    Google Scholar 

  • Hall RM, Vockler C (1987) The region of the IncN plasmid R46 coding for resistance to β-lactam antibiotics, streptomycin/spectinomycin and sulfonamides is closely related to antibiotic resistance segments found in IncW plasmids and in Tn21-like transposons. Nucleic Acids Res 15:7491–7501

    Google Scholar 

  • Heffron F (1983) Tn3 and its relatives. In: Shapiro J (ed) Mobile genetic elements. Academic Press, New York London: 223–260

    Google Scholar 

  • Hyde DR, Tu C-PD (1985) tnpM: A novel regulatory gene that enhances Tn21 transposition and suppresses cointegrate resolution. Cell 42:629–638

    Google Scholar 

  • Jenkins ST, Bennett PM (1978) Effect of mutations in deoxyribonucleic acid repair pathways on the sensitivity of Escherichia coli K-12 strains to nitrofurantoin. J Bacteriol 125:1214–1216

    Google Scholar 

  • Kratz J, Schmidt F, Wiedemann B (1983) Characterization of Tn2411 and Tn2410, two transposons derived from R-plasmid R1767 and related to Tn2603 and Tn21. J Bacteriol 155:1333–1342

    Google Scholar 

  • Kuhstoss S, Richardson MA, Rao N (1989) Site-specific integration in Streptomyces ambofaciens: localization of integration functions in S. ambofaciens plasmid pSAM2. J Bacteriol 171:16–23

    Google Scholar 

  • Lafond M, Couture F, Vezina G, Levesque RC (1989) Evolutionary perspectives on multiresistance β-lactamase transposons. J Bacteriol 171:6423–6429

    Google Scholar 

  • Martinez E, de la Cruz F (1988) Transposon Tn21 encodes a recA-independent site-specific integration system. Mol Gen Genet 211:320–325

    Google Scholar 

  • Mercier J, Lachapele J, Couture F, Lafond M, Vezina G, Boissinot M, Levesque RC (1990) Structural and functional characterization of tnpI, a recombinase locus in Tn21 and related β-lactamase transposon. J Bacteriol 172:3745–3757

    Google Scholar 

  • Nobuta K, Tolmanski ME, Crosa LM, Crosa JH (1988) Sequencing and expression of the 6′-N-acetyltransferase gene of transposon Tn1331 from Klebsiella pneumoniae. J Bacteriol 170:3769–3773

    Google Scholar 

  • Norrander J, Kempe T, Messing J (1983) Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene 26:101–106

    Google Scholar 

  • Novick RP, Clowes RC, Hohen SN, Curtiss III R, Datta N, Falkow S (1976) Uniform nomenclature of bacterial plasmids: a proposal. Bacteriol Rev 40:168–189

    Google Scholar 

  • Nücken EJ (1989) Molekulargenetische Untersuchungen zur “site spezifischen” Genintegration an den Enden des aadA-Gens in Tn21-Transposons. Ph. D. Thesis, University of Bonn, FRG

    Google Scholar 

  • Nücken EJ, Henschke RB, Schmidt FRJ (1989) DNA sequence of an OXA-2 β-lactamase gene from the R-factor R1767-derived plasmid pBP11 and comparison to other ampicillin resistance genes. J Gen Microbiol 135:761–765

    Google Scholar 

  • Olson ER, Chung S-T (1988) Transposon Tn4556 of Streptomyces fradiae: Nucleotide sequence of the ends and the target sites. J Bacteriol 170:1955–1957

    Google Scholar 

  • Ouellette M, Roy PH (1987) Homology of ORFs from Tn2603 and from R46 to site-specific recombinases. Nucleic Acids Res 15:10055

    Google Scholar 

  • Ouellette M, Bissonnette L, Roy PH (1987) Precise insertion of antibiotic resistance determinants into Tn21-like transposons: Nucleotide sequence of the OXA-I β-lactamase gene. Proc Natl Acad Sci USA 84:7378–7382

    Google Scholar 

  • Rüther U (1982) pUR250 allows rapid chemical sequencing of both DNA strands of its insert. Nucl Acids Res 10:5765–5772

    Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning, a laboratory manual. Cold Spring Harbor Laboratory, Press, Cold Spring Harbor, New York

    Google Scholar 

  • Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Nat Acad Sci USA 74:5463–5467

    Google Scholar 

  • Schmidt F (1984) The role of insertions, deletions, and substitutions in the evolution of R6 related plasmids encoding aminoglycoside transferase ANT(2″). Mol Gen Genet 194:248–259

    Google Scholar 

  • Schmidt F, Klopfer-Kaul I (1984) Evolutionary relationship between Tn21-like elements and pBP201, a plasmid from Klebsiella pneumoniae mediating resistance to gentamicin and eight other drugs. Mol Gen Genet 197:109–119

    Google Scholar 

  • Schmidt FRJ, Nücken EJ, Henschke RB (1988) Nucleotide sequence analysis of 2″-aminoglycoside nucleotidyl-transferase ANT(2″) from Tn4000: its relationship with AAD(3″) and impact on Tn21 evolution. Mol Microbiol 2:709–717

    Google Scholar 

  • Schmidt FRJ, Nücken EJ, Henschke RB (1989) Structure and function of hot spots providing signals for site-directed specific recombination and gene expression in Tn21 transposons. Mol Microbiol 3:1545–1555

    Google Scholar 

  • Schmidt FRJ, Henschke RB (1990) Mikrobielle Genetik, Ökologie und Sicherheitsforschung von Bodenmikroorganismen. In: Albrecht S (ed) Die Zukunft der Nutzpflanzen. Biotechnologie in Landwirtschaft und Pflanzenzuchtung. Campus Verlag, Frankfurt-New York, pp 127–148

    Google Scholar 

  • Schmitt R, Mötsch S, Rogowsky P, de la Cruz F, Grinsted J (1985) On the transposition and evolution of Tn1721 and its relatives. In: Helinksy DR, Cohen SN, Clewell DB, Jackson DA, Hollaender A (eds) Plasmids in bacteria. Plenum Press, New York, pp 79–91

    Google Scholar 

  • Scholz P, Haring V, Wittmann-Liebold B, Ashman K, Bagdasarian M, Scherzinger E (1989) Complete nucleotide sequence and gene organization of the broad-host-range plasmid RSF1010. Gene 75:271–288

    Google Scholar 

  • Shen P, Huang HV (1986) Homologous recombination in Escherichia coli: dependence on substrate length and homology. Genetics 112:441–457

    Google Scholar 

  • Simon R, Priefer U, Pühler A (1983) A broad host range mobilization system for in vivo genetic engineering transposon mutagenesis in gram-negative bacteria. BioTechnol Nov: 784–795

  • Stokes HW, Hall RM (1989) A novel family of potentially mobile DNA elements encoding site-specific gene-integration functions: integrons. Mol Microbiol 3 (12):1669–1683

    Google Scholar 

  • Sundström L, Radström P, Swedberg G, Sköld O (1988) Site-specific recombination promotes linkage between trimethoprim and sulfonamide resistance genes. Sequence characterization of dhfrV and sulI and a recombination active locus of Tn21. Mol Gen Genet 213:191–201

    Google Scholar 

  • Sundström L, Sköld O (1990) The dhfrI trimethoprim resistance gene of Tn7 can be found at specific sites in other genetic surroundings. Antimicro Agents Chemoter 34:642–650

    Google Scholar 

  • Watt VM, Ingles CJ, Urdea MS, Rutter W (1985) Homology requirements for recombination in Escherichia coli. Proc Natl Acad Sci USA 82:4768–4772

    Google Scholar 

  • Wohlleben W, Arnold W, Bissonnette L, Pelletier A, Tanguay A, Roy PH, Gamboa GC, Barry GF, Aubert E, Davies J, Kagan SA (1989) On the evolution of Tn21-like multiresistance transposons: Sequence analysis of the gene (aacC1) for gentamicin acetyltransferase-3-I, another member of the Tn21-based expression cassette. Mol Gen Genet 217:202–208

    Google Scholar 

  • Wood WB (1966) Host specificity of DNA produced by Escherichia coli: Bacterial mutations affecting the restriction and modification of DNA. J Mol Biol 169:118–123

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institut für Bodenbiologie, Bundesforschungsanstalt für Landwirtschaft (FAL), Bundesallee 50, W-3300, Braunschweig, Germany

    E. J. Nücken, R. B. Henschke & F. R. J. Schmidt

Authors
  1. E. J. Nücken
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. R. B. Henschke
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. F. R. J. Schmidt
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

Communicated by B.J. Kilbey

Rights and permissions

Reprints and permissions

About this article

Cite this article

Nücken, E.J., Henschke, R.B. & Schmidt, F.R.J. Site-specific integration of genes into hot spots for recombination flanking aadA in Tn21 transposons. Molec. Gen. Genet. 229, 137–146 (1991). https://doi.org/10.1007/BF00264222

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00264222

Key words

Search

Navigation