Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Nov;164(2):782–796. doi: 10.1128/jb.164.2.782-796.1985

Complete nucleotide sequence of macrolide-lincosamide-streptogramin B-resistance transposon Tn917 in Streptococcus faecalis.

J H Shaw, D B Clewell
PMCID: PMC214320  PMID: 2997130

Abstract

Streptococcus faecalis transposon Tn917 was cloned in Escherichia coli on plasmid vector pBR325. The erythromycin resistance determinant of Tn917 was not expressed in the E. coli background. The nucleotide sequence of Tn917 was determined and found to be 5,257 base pairs in length. Six open reading frames (ORFs) were identified and designated 1 through 6 (5' to 3'); all were on the same DNA strand. A region exhibiting strong homology with known promoters was identified upstream from ORF1. ORFs 1 to 3 were virtually identical to the previously sequenced erythromycin resistance determinant on Streptococcus sanguis plasmid pAM77. At the 3' point, where the homology between Tn917 and pAM77 ends, was a 20-base-pair region about 80% homologous with a component of the res site of Tn3. The amino acid sequence of ORF4 showed homology with other site-specific recombination enzymes, including approximately 30% homology with the resolvase of Tn3. Contained within Tn917 was a directly oriented 73-base-pair duplication of the left terminus. The Tn917 sequence revealed that antibiotic-enhanced transposition might be due to extension of transcription from the resistance-related genes (in ORFs 1 to 3) into transposition genes (in ORFs 4 to 6). Transcription analyses resulted in data consistent with this interpretation.

Full text

PDF
796

Images in this article

Selected References

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

  1. Banai M., LeBlanc D. J. Streptococcus faecalis R plasmid pJH1 contains an erythromycin resistance transposon (Tn3871) similar to transposon Tn917. J Bacteriol. 1984 Jun;158(3):1172–1174. doi: 10.1128/jb.158.3.1172-1174.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  3. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bolivar F. Construction and characterization of new cloning vehicles. III. Derivatives of plasmid pBR322 carrying unique Eco RI sites for selection of Eco RI generated recombinant DNA molecules. Gene. 1978 Oct;4(2):121–136. doi: 10.1016/0378-1119(78)90025-2. [DOI] [PubMed] [Google Scholar]
  5. Clewell D. B., Tomich P. K., Gawron-Burke M. C., Franke A. E., Yagi Y., An F. Y. Mapping of Streptococcus faecalis plasmids pAD1 and pAD2 and studies relating to transposition of Tn917. J Bacteriol. 1982 Dec;152(3):1220–1230. doi: 10.1128/jb.152.3.1220-1230.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Clewell D. B., Yagi Y., Bauer B. Plasmid-determined tetracycline resistance in Streptococcus faecalis: evidence for gene amplification during growth in presence of tetracycline. Proc Natl Acad Sci U S A. 1975 May;72(5):1720–1724. doi: 10.1073/pnas.72.5.1720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cooper T. G., Whitney P., Magasanik B. Reaction of lac-specific ribonucleic acid from Escherichia coli with lac deoxyribonucleic acid. J Biol Chem. 1974 Oct 25;249(20):6548–6555. [PubMed] [Google Scholar]
  8. Deininger P. L. Random subcloning of sonicated DNA: application to shotgun DNA sequence analysis. Anal Biochem. 1983 Feb 15;129(1):216–223. doi: 10.1016/0003-2697(83)90072-6. [DOI] [PubMed] [Google Scholar]
  9. Diver W. P., Grinsted J., Fritzinger D. C., Brown N. L., Altenbuchner J., Rogowsky P., Schmitt R. DNA sequences of and complementation by the tnpR genes of Tn21, Tn501 and Tn1721. Mol Gen Genet. 1983;191(2):189–193. doi: 10.1007/BF00334812. [DOI] [PubMed] [Google Scholar]
  10. Foster T. J. Plasmid-determined resistance to antimicrobial drugs and toxic metal ions in bacteria. Microbiol Rev. 1983 Sep;47(3):361–409. doi: 10.1128/mr.47.3.361-409.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gawron-Burke C., Clewell D. B. Regeneration of insertionally inactivated streptococcal DNA fragments after excision of transposon Tn916 in Escherichia coli: strategy for targeting and cloning of genes from gram-positive bacteria. J Bacteriol. 1984 Jul;159(1):214–221. doi: 10.1128/jb.159.1.214-221.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Greenblatt J., Li J. The nusA gene protein of Escherichia coli. Its identification and a demonstration that it interacts with the gene N transcription anti-termination protein of bacteriophage lambda. J Mol Biol. 1981 Mar 25;147(1):11–23. doi: 10.1016/0022-2836(81)90076-0. [DOI] [PubMed] [Google Scholar]
  13. Grindley N. D., Lauth M. R., Wells R. G., Wityk R. J., Salvo J. J., Reed R. R. Transposon-mediated site-specific recombination: identification of three binding sites for resolvase at the res sites of gamma delta and Tn3. Cell. 1982 Aug;30(1):19–27. doi: 10.1016/0092-8674(82)90007-1. [DOI] [PubMed] [Google Scholar]
  14. Gryczan T. J., Grandi G., Hahn J., Grandi R., Dubnau D. Conformational alteration of mRNA structure and the posttranscriptional regulation of erythromycin-induced drug resistance. Nucleic Acids Res. 1980 Dec 20;8(24):6081–6097. doi: 10.1093/nar/8.24.6081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gryczan T., Israeli-Reches M., Del Bue M., Dubnau D. DNA sequence and regulation of ermD, a macrolide-lincosamide-streptogramin B resistance element from Bacillus licheniformis. Mol Gen Genet. 1984;194(3):349–356. doi: 10.1007/BF00425543. [DOI] [PubMed] [Google Scholar]
  16. Hahn J., Grandi G., Gryczan T. J., Dubnau D. Translational attenuation of ermC: a deletion analysis. Mol Gen Genet. 1982;186(2):204–216. doi: 10.1007/BF00331851. [DOI] [PubMed] [Google Scholar]
  17. Heffron F., McCarthy B. J., Ohtsubo H., Ohtsubo E. DNA sequence analysis of the transposon Tn3: three genes and three sites involved in transposition of Tn3. Cell. 1979 Dec;18(4):1153–1163. doi: 10.1016/0092-8674(79)90228-9. [DOI] [PubMed] [Google Scholar]
  18. Horinouchi S., Byeon W. H., Weisblum B. A complex attenuator regulates inducible resistance to macrolides, lincosamides, and streptogramin type B antibiotics in Streptococcus sanguis. J Bacteriol. 1983 Jun;154(3):1252–1262. doi: 10.1128/jb.154.3.1252-1262.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Horinouchi S., Weisblum B. Posttranscriptional modification of mRNA conformation: mechanism that regulates erythromycin-induced resistance. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7079–7083. doi: 10.1073/pnas.77.12.7079. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Horinouchi S., Weisblum B. The control region for erythromycin resistance: free energy changes related to induction and mutation to constitutive expression. Mol Gen Genet. 1981;182(2):341–348. doi: 10.1007/BF00269681. [DOI] [PubMed] [Google Scholar]
  21. Ike Y., Clewell D. B. Genetic analysis of the pAD1 pheromone response in Streptococcus faecalis, using transposon Tn917 as an insertional mutagen. J Bacteriol. 1984 Jun;158(3):777–783. doi: 10.1128/jb.158.3.777-783.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ikemura T., Dahlberg J. E. Small ribonucleic acids of Escherichia coli. I. Characterization by polyacrylamide gel electrophoresis and fingerprint analysis. J Biol Chem. 1973 Jul 25;248(14):5024–5032. [PubMed] [Google Scholar]
  23. Khan S. A., Novick R. P. Terminal nucleotide sequences of Tn551, a transposon specifying erythromycin resistance in Staphylococcus aureus: homology with Tn3. Plasmid. 1980 Sep;4(2):148–154. doi: 10.1016/0147-619x(80)90004-9. [DOI] [PubMed] [Google Scholar]
  24. Kleckner N. Transposable elements in prokaryotes. Annu Rev Genet. 1981;15:341–404. doi: 10.1146/annurev.ge.15.120181.002013. [DOI] [PubMed] [Google Scholar]
  25. Lai C. J., Dahlberg J. E., Weisblum B. Structure of an inducibly methylatable nucleotide sequence in 23S ribosomal ribonucleic acid from erythromycin-resistant Staphylococcus aureus. Biochemistry. 1973 Jan 30;12(3):457–460. doi: 10.1021/bi00727a015. [DOI] [PubMed] [Google Scholar]
  26. Lai C. J., Weisblum B. Altered methylation of ribosomal RNA in an erythromycin-resistant strain of Staphylococcus aureus. Proc Natl Acad Sci U S A. 1971 Apr;68(4):856–860. doi: 10.1073/pnas.68.4.856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lai C. J., Weisblum B., Fahnestock S. R., Nomura M. Alteration of 23 S ribosomal RNA and erythromycin-induced resistance to lincomycin and spiramycin in Staphylococcus aureus. J Mol Biol. 1973 Feb 15;74(1):67–72. doi: 10.1016/0022-2836(73)90355-0. [DOI] [PubMed] [Google Scholar]
  28. Larson R., Messing J. Apple II software for M13 shotgun DNA sequencing. Nucleic Acids Res. 1982 Jan 11;10(1):39–49. doi: 10.1093/nar/10.1.39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Macrina F. L., Jones K. R., Wood P. H. Chimeric streptococcal plasmids and their use as molecular cloning vehicles in Streptococcus sanguis (Challis). J Bacteriol. 1980 Sep;143(3):1425–1435. doi: 10.1128/jb.143.3.1425-1435.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Macrina F. L., Tobian J. A., Jones K. R., Evans R. P., Clewell D. B. A cloning vector able to replicate in Escherichia coli and Streptococcus sanguis. Gene. 1982 Oct;19(3):345–353. doi: 10.1016/0378-1119(82)90025-7. [DOI] [PubMed] [Google Scholar]
  31. McLaughlin J. R., Murray C. L., Rabinowitz J. C. Unique features in the ribosome binding site sequence of the gram-positive Staphylococcus aureus beta-lactamase gene. J Biol Chem. 1981 Nov 10;256(21):11283–11291. [PubMed] [Google Scholar]
  32. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  33. Murphy E. Nucleotide sequence of ermA, a macrolide-lincosamide-streptogramin B determinant in Staphylococcus aureus. J Bacteriol. 1985 May;162(2):633–640. doi: 10.1128/jb.162.2.633-640.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Perkins J. B., Youngman P. J. A physical and functional analysis of Tn917, a Streptococcus transposon in the Tn3 family that functions in Bacillus. Plasmid. 1984 Sep;12(2):119–138. doi: 10.1016/0147-619x(84)90058-1. [DOI] [PubMed] [Google Scholar]
  35. Phillips S., Novick R. P. Tn554--a site-specific repressor-controlled transposon in Staphylococcus aureus. Nature. 1979 Mar 29;278(5703):476–478. doi: 10.1038/278476a0. [DOI] [PubMed] [Google Scholar]
  36. Rollins L. D., Lee L. N., LeBlanc D. J. Evidence for a disseminated erythromycin resistance determinant mediated by Tn917-like sequences among group D streptococci isolated from pigs, chickens, and humans. Antimicrob Agents Chemother. 1985 Apr;27(4):439–444. doi: 10.1128/aac.27.4.439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rosenberg M., Court D. Regulatory sequences involved in the promotion and termination of RNA transcription. Annu Rev Genet. 1979;13:319–353. doi: 10.1146/annurev.ge.13.120179.001535. [DOI] [PubMed] [Google Scholar]
  38. Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
  39. 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]
  40. Schöffl F., Arnold W., Pühler A., Altenbuchner J., Schmitt R. The tetracycline resistance transposons Tn1721 and Tn1771 have three 38-base-pair repeats and generate five-base-pair direct repeats. Mol Gen Genet. 1981;181(1):87–94. doi: 10.1007/BF00339010. [DOI] [PubMed] [Google Scholar]
  41. Shine J., Dalgarno L. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342–1346. doi: 10.1073/pnas.71.4.1342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Shivakumar A. G., Hahn J., Grandi G., Kozlov Y., Dubnau D. Posttranscriptional regulation of an erythromycin resistance protein specified by plasmic pE194. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3903–3907. doi: 10.1073/pnas.77.7.3903. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tinoco I., Jr, Borer P. N., Dengler B., Levin M. D., Uhlenbeck O. C., Crothers D. M., Bralla J. Improved estimation of secondary structure in ribonucleic acids. Nat New Biol. 1973 Nov 14;246(150):40–41. doi: 10.1038/newbio246040a0. [DOI] [PubMed] [Google Scholar]
  44. Tomich P. K., An F. Y., Clewell D. B. A transposon (Tn917) in Streptococcus faecalis that exhibits enhanced transposition during induction of drug resistance. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):1217–1221. doi: 10.1101/sqb.1979.043.01.138. [DOI] [PubMed] [Google Scholar]
  45. Tomich P. K., An F. Y., Clewell D. B. Properties of erythromycin-inducible transposon Tn917 in Streptococcus faecalis. J Bacteriol. 1980 Mar;141(3):1366–1374. doi: 10.1128/jb.141.3.1366-1374.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Tomich P. K., An F. Y., Damle S. P., Clewell D. B. Plasmid-related transmissibility and multiple drug resistance in Streptococcus faecalis subsp. zymogenes strain DS16. Antimicrob Agents Chemother. 1979 Jun;15(6):828–830. doi: 10.1128/aac.15.6.828. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Weisblum B., Graham M. Y., Gryczan T., Dubnau D. Plasmid copy number control: isolation and characterization of high-copy-number mutants of plasmid pE194. J Bacteriol. 1979 Jan;137(1):635–643. doi: 10.1128/jb.137.1.635-643.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wu A. M., Platt T. Transcription termination: nucleotide sequence at 3' end of tryptophan operon in Escherichia coli. Proc Natl Acad Sci U S A. 1978 Nov;75(11):5442–5446. doi: 10.1073/pnas.75.11.5442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Yagi Y., McLellan T. S., Frez W. A., Clewell D. B. Characterization of a small plasmid determining resistance to erythromycin, lincomycin, and vernamycin Balpha in a strain of Streptococcus sanguis isolated from dental plaque. Antimicrob Agents Chemother. 1978 May;13(5):884–887. doi: 10.1128/aac.13.5.884. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Youngman P. J., Perkins J. B., Losick R. Genetic transposition and insertional mutagenesis in Bacillus subtilis with Streptococcus faecalis transposon Tn917. Proc Natl Acad Sci U S A. 1983 Apr;80(8):2305–2309. doi: 10.1073/pnas.80.8.2305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Youngman P., Perkins J. B., Losick R. A novel method for the rapid cloning in Escherichia coli of Bacillus subtilis chromosomal DNA adjacent to Tn917 insertions. Mol Gen Genet. 1984;195(3):424–433. doi: 10.1007/BF00341443. [DOI] [PubMed] [Google Scholar]
  52. Youngman P., Perkins J. B., Losick R. Construction of a cloning site near one end of Tn917 into which foreign DNA may be inserted without affecting transposition in Bacillus subtilis or expression of the transposon-borne erm gene. Plasmid. 1984 Jul;12(1):1–9. doi: 10.1016/0147-619x(84)90061-1. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES