1887

Abstract

SUMMARY

To identify host genes that might influence nod (nodulation) gene expression in Rhizobium leguminosarum, a nodC-phoA reporter plasmid (carrying nodD) was introduced into a chemically mutagenized population of a R. leguminosarum strain lacking a symbiotic plasmid. The transconjugants were screened for expression of alkaline phosphatase (PhoA) on plates containing hesperetin, an inducer of nod genes, and a mutant with reduced expression was identified. When the nodC-phoA plasmid was cured from the mutant and the symbiotic plasmid pRL1jl introduced, the mutant formed nodules, but symbiotic nitrogen fixation was less than 20% of normal. When the nodC-phoA allele was introduced on pRL1jl a low level of nod gene induction was found. The reduced nodC expression appeared to be caused by a decrease in expression of the regulatory gene nodD, since expression of a nodD-lacZ fusion was also lower in the mutant than in the control. These mutant phenotypes and the low nitrogen fixation were complemented with a plasmid (plJ1848) from a R. leguminosarum cosmid library. DNA hybridization confirmed that plJ1848 was not from the symbiotic plasmid and showed that a DNA insertion was present in the mutant. The complementing region of plJ1848 was defined by transposon mutagenesis; DNA sequencing revealed that it carried the dicarboxylic acid transport (dct) genes. However, the mutant grew well with succinate as sole C-source. Genetic analysis revealed that the mutant appeared to contain IS50 in the regulatory gene dctB and that this mutation caused the reduction in nod gene expression. The effect was allele-specific since other mutations in dctB did not influence nod gene expression. Surprisingly, the mutant had a constitutive high level of succinate transport, indicating that the mutation caused unregulated expression of dctA the structural gene for dicarboxylic acid transport. This in some way appears to have lowered the expression of nodD, indicating that the nodD promoter may be influenced by the metabolic status of the cells or by expression of dctD in the absence of dctB.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-141-1-103
1995-01-01
2021-07-29
Loading full text...

Full text loading...

/deliver/fulltext/micro/141/1/mic-141-1-103.html?itemId=/content/journal/micro/10.1099/00221287-141-1-103&mimeType=html&fmt=ahah

References

  1. Baev N, Amar M, Defez R, laccarino M. 1992; The expression of the nodD and nodABC genes of Rhizobium leguminosarum is not regulated in response to combined nitrogen. . FEMS Microbiol Lett 97:205–208
    [Google Scholar]
  2. Beringer J. 1974; R factor transfer in Rhi^obium leguminosarum. l . J Gen Microbio 84:188–199
    [Google Scholar]
  3. Beringer J.E, wells B. 1977; The isolation of conditional ineffective mutants of Rhi^obium leguminosarum. l . J GenMicrobiol 98:339–343
    [Google Scholar]
  4. Buchanan-Wollaston v. 1979; Generalized transduction in Rhi^obium leguminosarum. l . J Gen Microbiol 112:135–142
    [Google Scholar]
  5. Burn J.E, Rossen L, Johnston A. W. B. 1987; Four classes of mutations in the nodD gene of Rhi^obium leguminosarum biovar viciae that affect its ability to autoregulate and/or activate other nod genes in the presence of flavonoid inducers. l . Genes & Dev 1:456–464
    [Google Scholar]
  6. Cubo T, Economou A, Economou G, Johnston G, Downie J.A. 1992; Molecular characterization and regulation of the rhizosphere-expressed genes rhiABCR that can influence nodulation by Rhhobium leguminosarum biovar viciae. l . f Bacteriol 174:4026–4035
    [Google Scholar]
  7. Djordjevic M. A., Redmond J. W., Batley M, Rolfe B. G. 1987; Clovers secrete specific phenolic compounds which either stimulate or repress nod gene expression in Rhi^obium trifolii. l . EMBOJ 6:1173–1179
    [Google Scholar]
  8. Downie J. A. 1991; A nod of recognition. l . Curr Biol 1:382–384
    [Google Scholar]
  9. Downie J. A., Knight C. D., Hombrecher G, Johnston A. W. B. 1993; Cloning of the symbiotic region of Rhi^obium leguminosarum-. the nodulation genes are between the nitrogenase genes and the nifAAike. gene l . EMBO J 2:947–952
    [Google Scholar]
  10. Dusha I, Kondorosi A. 1993; Genes at different regulatory levels are required for the ammonia control of nodulation in Rhi^obium meliloti . Mol & Gen Genet 240:435–444
    [Google Scholar]
  11. Economou A. 1990; Genetic analysis of a region of the Rhi^obium leguminosarum SymplasmidpRLIJI. PhD thesis, . University of East Anglia, UK
    [Google Scholar]
  12. Figurski D. H., Helinski D. R. 1979; Replication of an origin- containing derivative of plasmid RK2 dependent on a plasmid function provided in trans . Proc Natl Acad Sci USA 76:1648–1652
    [Google Scholar]
  13. Figurski D. H., Helinski D. R. 1979; Replication of an origin- containing derivative of plasmid RK2 dependent on a plasmid function provided in trans . Proc Natl Acad Sci USA 76:1648–1652
    [Google Scholar]
  14. Firmin J. L., Wilson K. E., Johnston A. .W. B. 1986; Flavonoid of nodulation genes in Rhi^obium reversed by other compounds present in plants. . Nature 324:90–92
    [Google Scholar]
  15. Gfittfert M. 1993; Regulation and function of rhizobial nodu- lation genes. . FEMS Microbiol Rev 104:39–64
    [Google Scholar]
  16. Hong G.-F., Burn J. E., Johnston A. .W. B. 1986; Evidence that DNA involved in the expression of nodulation nod genes in Rhi^obium, binds to the product of the regulatory gene nodD. . Nucleic Acids Res 15:9677–9690
    [Google Scholar]
  17. Jiang J., Albright L. M.., Nixon B. T. 1989; Conservation between coding and regulatory elements of Rhi^obium meliloti and Rhisobium leguminosarum dct genes. . J Bacteriol 171:5244–5253
    [Google Scholar]
  18. Johnston A. W. B, Beynon L. M.., Nixon J. L, Buchanan-Wollaston S. M., Setchell S. M, Hirsh P, Beringer J. E. 1978; High frequency transfer of nodulating ability between strains and species of Rhisobium. . Nature 276:634–636
    [Google Scholar]
  19. Kondorosi E., Gyuris J, Schmidt J., Buchanan-Wollaston S. M., Setchell S. M, John M, Duda E, Hoffmann B, Schell J, Kondorosi A. 1989; Positive and negative control of nod gene expression in Rhi^obium meliloti is required for optimal nodulation. . EMBO J 8:1331–1340
    [Google Scholar]
  20. Lamb J. W., Hombrecher G, Johnston A. W. B. 1982; Plasmid- determined nodulation and nitrogen-fixation abilities in Rhi^obium phaseoli. . Mol & Gen Genet 186:449–452
    [Google Scholar]
  21. Lamb J. W., Downie J. W., Johnston A. W. B. 1985; Cloning of the nodulation (nod) genes of Rhi^obium phaseoli and their homology to R. . leguminosarum nod DNA. Gene 34:235–241
    [Google Scholar]
  22. Maniatis T, Fritsch E. F., Sambrook J. 1982; Molecular Cloning: a Laboratory Manual. . Cold Spring Harbor, NY: Cold Spring Harbor Laboratory. Gene
    [Google Scholar]
  23. Poole P. S, Franklin M, Glenn A. R, Dilworth M. J. 1985; The transport of L-glutamate by Rhi^obium leguminosarum involves a common amino acid carrier. . J Gen Microbiol 131:1441–1448
    [Google Scholar]
  24. Recourt K, Schripsema J, Kijne J. W, van Brussel A, Lugtenberg B. J. J. 1991; Inoculation of Vicia sativa subsp nigra roots with Rhi^obium leguminosarum biovar viciae results in release of nod gene activating flavonoids and chalcones. . Riant Mol Biol 16:841–852
    [Google Scholar]
  25. Reznikoff W. S. 1993; The Tn 5 transposon. . Annu Rev Microbiol 47:945–963
    [Google Scholar]
  26. Ronson C. W., Astwood P. M. 1985; Genes involved in the carbon metabolism of bacteroids. In Nitrogen Fixation Research Progress, pp. 201-207. Edited by H. J. Evans, P. J. Bottomley & W. E. Newton. Dordrecht: Martinus Nijhoff. .
    [Google Scholar]
  27. Ronson C. W., Lyttleton P, Robertson J. G. 1981; dicarboxylate transport mutants in Rhi^obium trifolii form ineffective nodules on Trifolium repens. . Proc Natl Acad Sci USA 78:4284–4288
    [Google Scholar]
  28. Ronson C. W., Astwood P. M., Downie J. G. 1984; Molecular cloning and genetic organization of C4-dicarboxylate transport genes from R hi^obium leguminosarum. . J Bacteriol 160:903–909
    [Google Scholar]
  29. Ronson C. W., Astwood P. M., Nixon B. T, Ausubel F. M. 1987a; Deduced products of C4-dicarboxylate transport regu- latory genes of Rhi^obium leguminosarum are homologous to nitrogen regulatory products. . Nucleic Acids Res 15:7921–7934
    [Google Scholar]
  30. Ronson C. W., Nixon B. T, Albright L. M, Ausubel F. M. 1987b; Rhi^obium meliloti ntrA (rpoN ) gene is required for diverse metabolic functions. . J Bacteriol 169:2424–2431
    [Google Scholar]
  31. Rossen L, Shearman B. T, Albright C. A., Downie J. A. 1985; The nodD gene of Rhi^obium leguminosarum is autoregulatory and in the presence of plant exudate induces the nodABC genes. . EMBO J 4:3369–3373
    [Google Scholar]
  32. Ruvkun G. B., Ausubel F. M. 1981; A general method for site- directed mutagenesis in prokaryotes.. Nature 289:85–88
    [Google Scholar]
  33. Sanjuan J., Grob P., Gttttfert M., Hennecke H., Stacey G. 1994; NodW is essential for full expression of the common nodulation genes in Bradjrhi^obium japonicum . Mol Plant-Microbe Interact 7:364–369
    [Google Scholar]
  34. Schlaman H. R. M., Horvath B, Vijgenboom E, Hennecke H., Okker R. J. H., Lugtenberg B. J. J. 1991; Suppression of nodulation gene ex- pression in bacteroids of Rhiypbium leguminosarum biovar viciae. . J Bacteriol 173:4277–4287
    [Google Scholar]
  35. Schlaman H. R. M., Okker R. J. H., Lugtenberg B. J. J. 1992; Regulation of nodulation gene expression by NodD in rhizobia.. J Bacteriol 174:5177–5182
    [Google Scholar]
  36. Simon R., Quandt J., Klipp W. 1989; New derivatives of transposon Tn5 suitable for mobilization of replicons, generation of operon fusions and induction of genes in Gram-negative bacteria.. Gene 80:161–182
    [Google Scholar]
  37. Spaink H.P, Wijffelman C. A, Pees W, Okker H., Lugtenberg B. J. J. 1987; Rhi^obium nodulation gene nodD is a determinant of host specificity. . Nature 328:337–339
    [Google Scholar]
  38. Spaink H, Wijffelman C. A, Pees W, Okker R. J. H, Lugtenberg B. J. J. 1989; Localization of functional regions of the Rhi^obium nodD product using hybrid nodD . genes. 12:59–73
    [Google Scholar]
  39. Surin B. P, Watson J. M., Hamilton W. D. O, Okker R. J. H, Economou A, Downie J. A. 1990; Localization of functional regions of the Rhi^obium nodD product using hybrid nodD . Mol Microbiol 4:245–252
    [Google Scholar]
  40. Watson R. J, Rastogi V. K., Chan Y.-K. 1993; Aspartate transport in R Rhi^obium meliloti. . J Gen Microbiol 139:1315–1323
    [Google Scholar]
  41. Yarosh O, Charles T. C., Finan T. M. 1989; Analysis of C4- dicarboxylate transport genes in . Rhizobium meliloti Mol Microbiol 3:813–823
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-141-1-103
Loading
/content/journal/micro/10.1099/00221287-141-1-103
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error