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. 1988 Sep;170(9):4365–4372. doi: 10.1128/jb.170.9.4365-4372.1988

Cloning, characterization, and sequencing of an accessory gene regulator (agr) in Staphylococcus aureus.

H L Peng 1, R P Novick 1, B Kreiswirth 1, J Kornblum 1, P Schlievert 1
PMCID: PMC211451  PMID: 2457579

Abstract

We have previously identified a gene in Staphylococcus aureus, agr, whose activity is required for high-level post-exponential-phase expression of a series of secreted proteins. In this paper, we describe the cloning of this gene in Escherichia coli by using an inserted transposon (Tn551) as a cloning probe. The cloned gene, consisting of a 241-codon open reading frame containing the site of the transposon insertion, was recloned to an S. aureus vector, pSK265, and shown to be functional in S. aureus. Activity was evaluated by determinations of alpha-hemolysin, beta-hemolysin, and toxic shock syndrome toxin-1 production in early-stationary-phase cultures. The cloned gene showed considerable variation with respect to different exoproteins and different host strains compared with the chromosomal agr determinant; this variation could not be attributed to the higher copy number of the cloned gene and probably reflects inapparent subtleties of the regulatory system.

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Selected References

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  1. Chang S., Cohen S. N. High frequency transformation of Bacillus subtilis protoplasts by plasmid DNA. Mol Gen Genet. 1979 Jan 5;168(1):111–115. doi: 10.1007/BF00267940. [DOI] [PubMed] [Google Scholar]
  2. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  3. Clewell D. B., Helinski D. R. Supercoiled circular DNA-protein complex in Escherichia coli: purification and induced conversion to an opern circular DNA form. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1159–1166. doi: 10.1073/pnas.62.4.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Coleman G., Jakeman C. M., Martin N. Patterns of total extracellular protein secretion by a number of clinically isolated strains of Staphylococcus aureus. J Gen Microbiol. 1978 Jul;107(1):189–192. doi: 10.1099/00221287-107-1-189. [DOI] [PubMed] [Google Scholar]
  5. Horinouchi S., Weisblum B. Nucleotide sequence and functional map of pC194, a plasmid that specifies inducible chloramphenicol resistance. J Bacteriol. 1982 May;150(2):815–825. doi: 10.1128/jb.150.2.815-825.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Jones C. L., Khan S. A. Nucleotide sequence of the enterotoxin B gene from Staphylococcus aureus. J Bacteriol. 1986 Apr;166(1):29–33. doi: 10.1128/jb.166.1.29-33.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Katsuno S., Kondo M. Regulation of staphylococcal enterotoxin B synthesis and its relation to other extracellular proteins. Jpn J Med Sci Biol. 1973 Feb;26(1):26–29. [PubMed] [Google Scholar]
  8. Kreiswirth B. N., Löfdahl S., Betley M. J., O'Reilly M., Schlievert P. M., Bergdoll M. S., Novick R. P. The toxic shock syndrome exotoxin structural gene is not detectably transmitted by a prophage. Nature. 1983 Oct 20;305(5936):709–712. doi: 10.1038/305709a0. [DOI] [PubMed] [Google Scholar]
  9. Mallonee D. H., Glatz B. A., Pattee P. A. Chromosomal mapping of a gene affecting enterotoxin A production in Staphylococcus aureus. Appl Environ Microbiol. 1982 Feb;43(2):397–402. doi: 10.1128/aem.43.2.397-402.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Morfeldt E., Janzon L., Arvidson S., Löfdahl S. Cloning of a chromosomal locus (exp) which regulates the expression of several exoprotein genes in Staphylococcus aureus. Mol Gen Genet. 1988 Mar;211(3):435–440. doi: 10.1007/BF00425697. [DOI] [PubMed] [Google Scholar]
  11. NOVICK R. P., RICHMOND M. H. NATURE AND INTERACTIONS OF THE GENETIC ELEMENTS GOVERNING PENICILLINASE SYNTHESIS IN STAPHYLOCOCCUS AUREUS. J Bacteriol. 1965 Aug;90:467–480. doi: 10.1128/jb.90.2.467-480.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Novick R. P., Adler G. K., Projan S. J., Carleton S., Highlander S. K., Gruss A., Khan S. A., Iordanescu S. Control of pT181 replication I. The pT181 copy control function acts by inhibiting the synthesis of a replication protein. EMBO J. 1984 Oct;3(10):2399–2405. doi: 10.1002/j.1460-2075.1984.tb02146.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Novick R. P., Brodsky R. Studies on plasmid replication. I. Plasmid incompatibility and establishment in Staphylococcus aureus. J Mol Biol. 1972 Jul 21;68(2):285–302. doi: 10.1016/0022-2836(72)90214-8. [DOI] [PubMed] [Google Scholar]
  14. Novick R. P., Edelman I., Schwesinger M. D., Gruss A. D., Swanson E. C., Pattee P. A. Genetic translocation in Staphylococcus aureus. Proc Natl Acad Sci U S A. 1979 Jan;76(1):400–404. doi: 10.1073/pnas.76.1.400. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Novick R. P., Murphy E., Gryczan T. J., Baron E., Edelman I. Penicillinase plasmids of Staphylococcus aureus: restriction-deletion maps. Plasmid. 1979 Jan;2(1):109–129. doi: 10.1016/0147-619x(79)90010-6. [DOI] [PubMed] [Google Scholar]
  16. Novick R. Properties of a cryptic high-frequency transducing phage in Staphylococcus aureus. Virology. 1967 Sep;33(1):155–166. doi: 10.1016/0042-6822(67)90105-5. [DOI] [PubMed] [Google Scholar]
  17. O'Callaghan C. H., Morris A., Kirby S. M., Shingler A. H. Novel method for detection of beta-lactamases by using a chromogenic cephalosporin substrate. Antimicrob Agents Chemother. 1972 Apr;1(4):283–288. doi: 10.1128/aac.1.4.283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. O'Reilly M., de Azavedo J. C., Kennedy S., Foster T. J. Inactivation of the alpha-haemolysin gene of Staphylococcus aureus 8325-4 by site-directed mutagenesis and studies on the expression of its haemolysins. Microb Pathog. 1986 Apr;1(2):125–138. doi: 10.1016/0882-4010(86)90015-x. [DOI] [PubMed] [Google Scholar]
  19. Projan S. J., Carleton S., Novick R. P. Determination of plasmid copy number by fluorescence densitometry. Plasmid. 1983 Mar;9(2):182–190. doi: 10.1016/0147-619x(83)90019-7. [DOI] [PubMed] [Google Scholar]
  20. Raibaud O., Schwartz M. Positive control of transcription initiation in bacteria. Annu Rev Genet. 1984;18:173–206. doi: 10.1146/annurev.ge.18.120184.001133. [DOI] [PubMed] [Google Scholar]
  21. Recsei P., Kreiswirth B., O'Reilly M., Schlievert P., Gruss A., Novick R. P. Regulation of exoprotein gene expression in Staphylococcus aureus by agar. Mol Gen Genet. 1986 Jan;202(1):58–61. doi: 10.1007/BF00330517. [DOI] [PubMed] [Google Scholar]
  22. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. 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]
  25. Schlievert P. M., Shands K. N., Dan B. B., Schmid G. P., Nishimura R. D. Identification and characterization of an exotoxin from Staphylococcus aureus associated with toxic-shock syndrome. J Infect Dis. 1981 Apr;143(4):509–516. doi: 10.1093/infdis/143.4.509. [DOI] [PubMed] [Google Scholar]
  26. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  27. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  28. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wang P. Z., Novick R. P. Nucleotide sequence and expression of the beta-lactamase gene from Staphylococcus aureus plasmid pI258 in Escherichia coli, Bacillus subtilis, and Staphylococcus aureus. J Bacteriol. 1987 Apr;169(4):1763–1766. doi: 10.1128/jb.169.4.1763-1766.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Yoshikawa M., Matsuda F., Naka M., Murofushi E., Tsunematsu Y. Pleiotropic alteration of activities of several toxins and enzymes in mutants of Staphylococcus aureus. J Bacteriol. 1974 Jul;119(1):117–122. doi: 10.1128/jb.119.1.117-122.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]

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