Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1994 Sep;176(18):5697–5703. doi: 10.1128/jb.176.18.5697-5703.1994

Inhibition of Agrobacterium tumefaciens oncogenicity by the osa gene of pSa.

C Y Chen 1, C I Kado 1
PMCID: PMC196773  PMID: 8083162

Abstract

The IncW plasmid pSa originally derived from Shigella flexneri completely inhibits the tumor-inducing ability of Agrobacterium tumefaciens when it is resident in this organism. Oncogenic inhibition is mediated through the expression of the osa gene on pSa. This gene is part of a 3.1-kb DNA segment of pSa that contains four open reading frames revealed by sequencing. Specific deletions and TnCAT insertions within this segment localized the oncogenic inhibitory activity to the last open reading frame, orf-4, designated osa (for oncogenic suppression activity). No promoter exists immediately upstream of the coding sequence of osa since TnCAT insertions or deletions into orf-3 caused the loss of oncogenic inhibition. Deletion analysis showed that the promoter of orf-1 is required for osa transcription. The first three orfs have no role in oncogenic inhibition, since osa alone placed under the control of a constitutive Pkm promoter completely inhibited A. tumefaciens oncogenicity. This inhibition of oncogenicity by osa is not limited to a specific host plant but appears to show broad host specificity. Because the osa-encoded product has close homologies to the fiwA-encoded product of the IncP plasmid RP1, osa may be involved in fertility inhibition that would prevent or reduce the formation of stable mating pairs and T-DNA transfer between A. tumefaciens and plants.

Full text

PDF
5697

Images in this article

Selected References

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

  1. Alton N. K., Vapnek D. Nucleotide sequence analysis of the chloramphenicol resistance transposon Tn9. Nature. 1979 Dec 20;282(5741):864–869. doi: 10.1038/282864a0. [DOI] [PubMed] [Google Scholar]
  2. Bolland S., Llosa M., Avila P., de la Cruz F. General organization of the conjugal transfer genes of the IncW plasmid R388 and interactions between R388 and IncN and IncP plasmids. J Bacteriol. 1990 Oct;172(10):5795–5802. doi: 10.1128/jb.172.10.5795-5802.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Close S. M., Kado C. I. A gene near the plasmid pSa origin of replication encodes a nuclease. Mol Microbiol. 1992 Feb;6(4):521–527. doi: 10.1111/j.1365-2958.1992.tb01497.x. [DOI] [PubMed] [Google Scholar]
  4. Close S. M., Kado C. I. The osa gene of pSa encodes a 21.1-kilodalton protein that suppresses Agrobacterium tumefaciens oncogenicity. J Bacteriol. 1991 Sep;173(17):5449–5456. doi: 10.1128/jb.173.17.5449-5456.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cooley M. B., D'Souza M. R., Kado C. I. The virC and virD operons of the Agrobacterium Ti plasmid are regulated by the ros chromosomal gene: analysis of the cloned ros gene. J Bacteriol. 1991 Apr;173(8):2608–2616. doi: 10.1128/jb.173.8.2608-2616.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Filichkin S. A., Gelvin S. B. Formation of a putative relaxation intermediate during T-DNA processing directed by the Agrobacterium tumefaciens VirD1,D2 endonuclease. Mol Microbiol. 1993 May;8(5):915–926. doi: 10.1111/j.1365-2958.1993.tb01637.x. [DOI] [PubMed] [Google Scholar]
  7. Fong S. T., Stanisich V. A. Location and characterization of two functions on RP1 that inhibit the fertility of the IncW plasmid R388. J Gen Microbiol. 1989 Mar;135(3):499–502. doi: 10.1099/00221287-135-3-499. [DOI] [PubMed] [Google Scholar]
  8. Gallie D. R., Zaitlin D., Perry K. L., Kado C. I. Characterization of the replication and stability regions of Agrobacterium tumefaciens plasmid pTAR. J Bacteriol. 1984 Mar;157(3):739–745. doi: 10.1128/jb.157.3.739-745.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Jasper F., Koncz C., Schell J., Steinbiss H. H. Agrobacterium T-strand production in vitro: sequence-specific cleavage and 5' protection of single-stranded DNA templates by purified VirD2 protein. Proc Natl Acad Sci U S A. 1994 Jan 18;91(2):694–698. doi: 10.1073/pnas.91.2.694. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kado C. I., Liu S. T. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol. 1981 Mar;145(3):1365–1373. doi: 10.1128/jb.145.3.1365-1373.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kado C. I. Promiscuous DNA transfer system of Agrobacterium tumefaciens: role of the virB operon in sex pilus assembly and synthesis. Mol Microbiol. 1994 Apr;12(1):17–22. doi: 10.1111/j.1365-2958.1994.tb00990.x. [DOI] [PubMed] [Google Scholar]
  12. Kamoun S., Cooley M. B., Rogowsky P. M., Kado C. I. Two chromosomal loci involved in production of exopolysaccharide in Agrobacterium tumefaciens. J Bacteriol. 1989 Mar;171(3):1755–1759. doi: 10.1128/jb.171.3.1755-1759.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kao J. C., Perry K. L., Kado C. I. Indoleacetic acid complementation and its relation to host range specifying genes on the Ti plasmid of Agrobacterium tumefaciens. Mol Gen Genet. 1982;188(3):425–432. doi: 10.1007/BF00330044. [DOI] [PubMed] [Google Scholar]
  14. Klapwijk P. M., van Beelen P., Schilperoort R. A. Isolation of a recombination deficient Agrobacterium tumefaciens mutant. Mol Gen Genet. 1979 Jun 7;173(2):171–175. doi: 10.1007/BF00330307. [DOI] [PubMed] [Google Scholar]
  15. Lessl M., Balzer D., Pansegrau W., Lanka E. Sequence similarities between the RP4 Tra2 and the Ti VirB region strongly support the conjugation model for T-DNA transfer. J Biol Chem. 1992 Oct 5;267(28):20471–20480. [PubMed] [Google Scholar]
  16. Lessl M., Pansegrau W., Lanka E. Relationship of DNA-transfer-systems: essential transfer factors of plasmids RP4, Ti and F share common sequences. Nucleic Acids Res. 1992 Nov 25;20(22):6099–6100. doi: 10.1093/nar/20.22.6099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Loper J. E., Kado C. I. Host range conferred by the virulence-specifying plasmid of Agrobacterium tumefaciens. J Bacteriol. 1979 Aug;139(2):591–596. doi: 10.1128/jb.139.2.591-596.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Matthysse A. G. Effect of Plasmid pSa and of Auxin on Attachment of Agrobacterium tumefaciens to Carrot Cells. Appl Environ Microbiol. 1987 Oct;53(10):2574–2582. doi: 10.1128/aem.53.10.2574-2582.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Morrison D. A. Transformation and preservation of competent bacterial cells by freezing. Methods Enzymol. 1979;68:326–331. doi: 10.1016/0076-6879(79)68023-0. [DOI] [PubMed] [Google Scholar]
  20. Okumura M. S., Kado C. I. The region essential for efficient autonomous replication of pSa in Escherichia coli. Mol Gen Genet. 1992 Oct;235(1):55–63. doi: 10.1007/BF00286181. [DOI] [PubMed] [Google Scholar]
  21. Pansegrau W., Schoumacher F., Hohn B., Lanka E. Site-specific cleavage and joining of single-stranded DNA by VirD2 protein of Agrobacterium tumefaciens Ti plasmids: analogy to bacterial conjugation. Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11538–11542. doi: 10.1073/pnas.90.24.11538. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pohlman R. F., Liu F., Wang L., Moré M. I., Winans S. C. Genetic and biochemical analysis of an endonuclease encoded by the IncN plasmid pKM101. Nucleic Acids Res. 1993 Oct 25;21(21):4867–4872. doi: 10.1093/nar/21.21.4867. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Shirasu K., Kado C. I. Membrane location of the Ti plasmid VirB proteins involved in the biosynthesis of a pilin-like conjugative structure on Agrobacterium tumefaciens. FEMS Microbiol Lett. 1993 Aug 1;111(2-3):287–294. doi: 10.1111/j.1574-6968.1993.tb06400.x. [DOI] [PubMed] [Google Scholar]
  25. Shirasu K., Kado C. I. The virB operon of the Agrobacterium tumefaciens virulence regulon has sequence similarities to B, C and D open reading frames downstream of the pertussis toxin-operon and to the DNA transfer-operons of broad-host-range conjugative plasmids. Nucleic Acids Res. 1993 Jan 25;21(2):353–354. doi: 10.1093/nar/21.2.353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Shirasu K., Koukolíková-Nicola Z., Hohn B., Kado C. I. An inner-membrane-associated virulence protein essential for T-DNA transfer from Agrobacterium tumefaciens to plants exhibits ATPase activity and similarities to conjugative transfer genes. Mol Microbiol. 1994 Feb;11(3):581–588. doi: 10.1111/j.1365-2958.1994.tb00338.x. [DOI] [PubMed] [Google Scholar]
  27. Tait R. C., Lundquist R. C., Kado C. I. Genetic map of the crown gall suppressive IncW plasmid pSa. Mol Gen Genet. 1982;186(1):10–15. doi: 10.1007/BF00422905. [DOI] [PubMed] [Google Scholar]
  28. Tait R. C., Rempel H., Rodriguez R. L., Kado C. I. The aminoglycoside-resistance operon of the plasmid pSa: nucleotide sequence of the streptomycin-spectinomycin resistance gene. Gene. 1985;36(1-2):97–104. doi: 10.1016/0378-1119(85)90073-3. [DOI] [PubMed] [Google Scholar]
  29. Watanabe T., Furuse C., Sakaizumi S. Transduction of various R factors by phage P1 in Escherichia coli and by phage P22 in Salmonella typhimurium. J Bacteriol. 1968 Nov;96(5):1791–1795. doi: 10.1128/jb.96.5.1791-1795.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Waters V. L., Strack B., Pansegrau W., Lanka E., Guiney D. G. Mutational analysis of essential IncP alpha plasmid transfer genes traF and traG and involvement of traF in phage sensitivity. J Bacteriol. 1992 Oct;174(20):6666–6673. doi: 10.1128/jb.174.20.6666-6673.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Winans S. C., Walker G. C. Fertility inhibition of RP1 by IncN plasmid pKM101. J Bacteriol. 1985 Jan;161(1):425–427. doi: 10.1128/jb.161.1.425-427.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Yusoff K., Stanisich V. A. Location of a function on RP1 that fertility inhibits Inc W plasmids. Plasmid. 1984 Mar;11(2):178–181. doi: 10.1016/0147-619x(84)90022-2. [DOI] [PubMed] [Google Scholar]

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

RESOURCES