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In Vivo Expression Technology (IVET) and Its Application in Plant-Associated Bacteria

  • Lee, Seon-Woo (Agrochemical Research Team, Korea Research Institute of Chemical Technology(KRICT))
  • Published : 2002.04.01

Abstract

In vivo expression technology (IVET) has been developed to study bacterial gene expression in Salmonella typhimurium during host infection. The expression of selected genes by IVET has been elevated in vivo but not in vitro. The selected genes turned out to be important for bacterial virulence and/or pathogenicity. IVET depends on a synthetic operon with a promoterless transcriptional fusion between a selection marker gene and a reporter gene. The IVET approach has been successfully adapted in other bacterial pathogens and plant-associated bacteria using different selection markers. Pseudomonas putida suppresses citrus root rot caused by Phytophthora parasitica and enhances citrus seedling growth. The WET strategy was adapted based on a transcriptional fusion, pyrBC'-lacZ, in P. putida to study the bacterial traits important far biocontrol activities. Several genes appeared to be induced on P. parasitica hyphae and were found to be related with metabolism and regulation of gene expression. It is likely that the biocontrol strain took a metabolic advantage from the plant pathogenic fungus and then suppressed citrus root rot effectively. The result was parallel with those from the adaptation of IVET in P. fluorescens, a plant growth promoting rhizobacteria (PGPR). Interestingly, genes encoding components for type III secretion system have been identified as rhizosphere-induced genes in the PGPR strain. The type III secretion system may play a certain role during interaction with its counterpart plants. Application of IVET has been demonstrated in a wide range of bacteria. It is an important strategy to genetically understand complicated bacterial traits in the environment.

Keywords

References

  1. Andersen, G. A., Beattie, G. A. and Lindow, S. E. 1998. Molecular characterization and sequence of a methionine biosynthetic locus from Pseudomonas syringae. J. Bacteriol. 180:4497-4507
  2. Autret, N., Dubail, Trieu-Cuot, P., Berche, P. and Charbit, A 2001. Identification of new genes involved in the virulence of Listeria monocytogenes by signature-tagged transposon mutagenesis, Infect. Immun. 69:2054-2065 https://doi.org/10.1128/IAI.69.4.2054-2065.2001
  3. Camilli, A. and Mekalanos, J. J. 1995. Use of recombinase gene fusions to identify Vibrio cholerae genes induced during infection. Mol. Microbiol. 18:671-683 https://doi.org/10.1111/j.1365-2958.1995.mmi_18040671.x
  4. Cook, R. J. 1993. Making greater use of introduced microorganisms for biological control of plant pathogens. Annu. Rev. Phytopathol. 31:53-80 https://doi.org/10.1146/annurev.py.31.090193.000413
  5. De Saizieu, A., Certa, U., Warrington, J., Gray, C., Keck, W. and Mous, J. 1998. Bacterial transcript imaging by hybridization of total RNA to oligonucleotide arrays. Nature Biotech. 16:45-48
  6. Handelsman, J. and Stabb. E. V. 1996. Biocontrol of soilborne plant pathogens. Plant Cell 8:1855-1869 https://doi.org/10.1105/tpc.8.10.1855
  7. Handfield, M., Schweizer, H. P., Mahan, M. J., Sanschagrin, F., Hoang, T. and Levesque, R. C. 1998. ASD-GFP vectors for in vivo expression technology in Pseudomonas aeruginosa and other Gram-negative bacteria. BioTechniques 24:261-264
  8. Heithoff; D. M., Conner, C. P., Hanna, P. C., Julio, S. M., Hentschel, T. and Mahan, M. J. 1997. Bacterial infection as assessed by in vivo gene expression. Proc. Natl. Acad. Sci. USA 94:934-939 https://doi.org/10.1073/pnas.94.3.934
  9. Heithoff, D. M., Sinsheimer, R. L., Low, D. A. and Mahan, M. J. 1999. An essential role for DNA adenine methylation in bacterial virulence. Science 284:96-970 https://doi.org/10.1126/science.284.5411.96
  10. Hensel, M. 1998. Whole genome scan of habitat genes by signature-tagged mutagenesis. Electrophoresis 19:608-612 https://doi.org/10.1002/elps.1150190425
  11. Hensel, M., Shea, J. E., Gleeson, C., Jones, M. D., Dalton, E. and Holden, D. 1995. Simultaneous identification of bacterial virulence genes by negative selection. Science 269:400-403 https://doi.org/10.1126/science.7618105
  12. Lai, Y. C., Peng, H. L. and Chang, H. Y. 2001. Identification of genes induced in vivo during Klebsiella pneumonias CG43 infection. Infect. Immun. 69:7140-7145 https://doi.org/10.1128/IAI.69.11.7140-7145.2001
  13. Lee, S.-W. and Cooksey, D. A. 2000. Genes expressed in Pseudomonas putida during colonization of a plant-pathogenic fungi. Appl. Environ. Microbiol. 66:2764-2772 https://doi.org/10.1128/AEM.66.7.2764-2772.2000
  14. Lee, S.-W., Glickmann, E. and Cooksey, D. A. 2001. Chromosomal locus for cadmium resistance in Pseudomonas putida consisting of a cadium-transporting ATPase and a MerR family response regulator. Appl. Environ. Microbiol. 67:1437-1444 https://doi.org/10.1128/AEM.67.4.1437-1444.2001
  15. Lee, S.-W., Menge, J. A. and Cooksey, D. A. 1998. Cloning genes expressed during colonization of fungal hyphae or citrus root tips by Pseudomonas putida. Phytopathology 88:S52 https://doi.org/10.1094/PHYTO.1998.88.1.52
  16. Lowe, A. M., Beattie, D. T. and Deresiewicz, R. L. 1998. Identification of novel staphylococcal virulence genes by in vivo expression technology. Mol. Microbiol. 27:967-976 https://doi.org/10.1046/j.1365-2958.1998.00741.x
  17. Mahan, M. J. and Low, D. A. 2001. DNA methylation regulates bacterial gene expression and virulence. ASM News 67:356-361
  18. Mahan, M. J., Slauch, J. M. and Mekalanos, J. J. 1993a. Bacteriophage P22 transduction of integrated plasmids: single-step cloning of Salmonella typhimurium gene fusions. J. Bacteriol. 175:7086-7091 https://doi.org/10.1128/jb.175.21.7086-7091.1993
  19. Mahan, M. J., Slauch, J. M. and Mekalanos, J. J. 1993b. Selection of virulence genes that are specifically induced in host tissues. Science 259:686-688 https://doi.org/10.1126/science.8430319
  20. Mahan, M. J., Tobias, J. W., Slauch, J. M., Hanna, P. C., Collier, R. J. and Mekalanos, J. J. 1995. Antibiotic-based selection for bacterial genes that are specifically induced during infection of a host. Proc. Natl. Acad. Sci. USA 92:669-673 https://doi.org/10.1073/pnas.92.3.669
  21. OFlaherty, S., Moenne-Loccoz, Y., Boesten, B., Higgins, P., Dowling, D. N., Condon, S. and OGara, F. 1995. Green house and field evaluations of an autoselective system based on an essential thymidylate synthase gene for improved maintenance of plasmid vectors in modified Rhizobium meliloti. Appl. Environ. Microbiol. 61:4051-4056
  22. Polesky, A. H., Ross, J. T., Falkow, S. and Tompkins, L. S. 2001. Identification of Legionella pneumophila genes important for infection of amoebas by signature-tagged mutagenesis. Infect. Immun. 69:977-987 https://doi.org/10.1128/IAI.69.2.977-987.2001
  23. Preston, G. M., Bertrand, N. and Rainey, P. B. 2001. Type III secretion in plant growth-promoting Pseudomonas fluorescens SBW25. Mol. Microbiol. 41:999-1014 https://doi.org/10.1046/j.1365-2958.2001.02560.x
  24. Rainey, P. B. 1999. Adaptation of Pseudomonas fluorescens to the plant rhizosphere. Environ. Microbiol. 1:243-257 https://doi.org/10.1046/j.1462-2920.1999.00040.x
  25. Ramsay, G. 1988. DNA chips: state-of-the-art. Nature Biotech. 16: 40-44 https://doi.org/10.1038/nbt0198-40
  26. Ross, P., OGara, F. and Codon, S. 1990. Thymidylate synthase gene from Lactococcus lactis as a genetic marker: an alternative to antibiotic resistance genes. Appl. Environ. Microbiol. 52:2164-2169
  27. Schena, M., Shalon, D., Davis, R. W. and Brown, P. O. 1995. Quantitative monitoring of gene expression patterns with a complementary DNA microarray. Science 270:467-470 https://doi.org/10.1126/science.270.5235.467
  28. Schurr, M. J., Vickrey, J. F., Kumar, A. P., Campbell, A. L., Cunin, R., Benjamin, R. C., Shanley, M. S. and ODonovan, G. A., 1995. Aspartate transcarbamoylase gene of Pseudomonas putida: requirement for an inactive dihyroorotase for assembly into the dodecameric holoenzyme. J. Bacteriol. 177:1751-1759 https://doi.org/10.1128/jb.177.7.1751-1759.1995
  29. Shea, J. E., Santangelo, J. D. and Feldman, R. G. 2000. Signaturetagged mutagenesis in the identification of virulence genes in pathogens. Curr. Opin. Microbiol. 3:451-458 https://doi.org/10.1016/S1369-5274(00)00120-X
  30. Slauch, J. M. and Camilli, A. 2000. IVET and RIVET: use of gene fusions to identify bacterial virulence factors specifically induced in host tissues. Methods Enzymol. 326:73-96 https://doi.org/10.1016/S0076-6879(00)26047-3
  31. Slauch, J. M. and Silhavy, T. J. 1991. Genetic fusions as experimental tools. Methods Enzymol. 204:213-248 https://doi.org/10.1016/0076-6879(91)04011-C
  32. Steddom, K. C. and Menge, J. A. 1999. Continuous application of the biocontrol bacterium, Pseudomonas putida 06909, improves biocontrol of Phytophthora parasitica on citrus. Phytopathology 89:S75
  33. Tumey, J. K. 1995. The biological control of Phytophthora root rot of citrus using rhizobacteria. Ph. D. thesis. University of California, Riverside
  34. Yang, C.-H., Menge, J. A. and Cooksey, D. A. 1994. Mutations affecting hyphal colonization and pyoverdine production in pseudomonads antagonistic toward Phytophthora parasitica. Appl. Environ. Microbiol. 60:473-481
  35. Young, G. M. and Miller, V. L. 1997. Identification of novel chromosomal loci affecting Yersinia enterolitica pathogenesis. Mol. Microbiol. 25: 319-328 https://doi.org/10.1046/j.1365-2958.1997.4661829.x
  36. Wang, J., Mushegian, A., Lory, S. and Jin, S. 1996. Large-scale isolation of candidate virulence genes of Pseudomonas aeruginosa by in vivo selection. Proc. Natl. Acad. Sci. USA 93: 10434-10439 https://doi.org/10.1073/pnas.93.19.10434
  37. Weller, D. M. 1988. Biological control of soilborne plant pathogens in the rhizosphere with bacterial. Annu. Rev. Phytopathol. 26:379-407 https://doi.org/10.1146/annurev.py.26.090188.002115

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