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. 1998 Jun;149(2):785–794. doi: 10.1093/genetics/149.2.785

Multiple recombination events maintain sequence identity among members of the nitrogenase multigene family in Rhizobium etli.

C Rodríguez 1, D Romero 1
PMCID: PMC1460202  PMID: 9611191

Abstract

A distinctive characteristic of the Rhizobium genome is the frequent finding of reiterated sequences, which often constitute multigene families. Interestingly, these families usually maintain a high degree of nucleotide sequence identity. It is commonly assumed that apparent gene conversion between reiterated elements might lead to concerted variation among members of a multigene family. However, the operation of this mechanism has not yet been demonstrated in the Rhizobiaceae. In this work, we employed different genetic constructions to address the role of apparent gene conversion as a homogenizing mechanism between members of the plasmid-located nitrogenase multigene family in Rhizobium etli. Our results show that a 28-bp insertion into one of the nitrogenase reiterations can be corrected by multiple recombination events, including apparent gene conversion. The correction process was dependent on the presence of both a wild-type recA gene and wild-type copies of the nitrogenase reiterations. Frequencies of apparent gene conversion to the wild-type nitrogenase reiterations were the same when the insertion to be corrected was located either in cis or in trans, indicating that this event frequently occurs through intermolecular interactions. Interestingly, a high frequency of multiple crossovers was observed, suggesting that these large plasmid molecules are engaging repeatedly in recombination events, in a situation akin to phage recombination or recombination among small, high-copy number plasmids.

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Selected References

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

  1. Abdulkarim F., Hughes D. Homologous recombination between the tuf genes of Salmonella typhimurium. J Mol Biol. 1996 Jul 26;260(4):506–522. doi: 10.1006/jmbi.1996.0418. [DOI] [PubMed] [Google Scholar]
  2. Badenoch-Jones J., Holton T. A., Morrison C. M., Scott K. F., Shine J. Structural and functional analysis of nitrogenase genes from the broad-host-range Rhizobium strain ANU240. Gene. 1989 Apr 15;77(1):141–153. doi: 10.1016/0378-1119(89)90368-5. [DOI] [PubMed] [Google Scholar]
  3. Bergman K., Nulty E., Su L. H. Mutations in the two flagellin genes of Rhizobium meliloti. J Bacteriol. 1991 Jun;173(12):3716–3723. doi: 10.1128/jb.173.12.3716-3723.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blattner F. R., Plunkett G., 3rd, Bloch C. A., Perna N. T., Burland V., Riley M., Collado-Vides J., Glasner J. D., Rode C. K., Mayhew G. F. The complete genome sequence of Escherichia coli K-12. Science. 1997 Sep 5;277(5331):1453–1462. doi: 10.1126/science.277.5331.1453. [DOI] [PubMed] [Google Scholar]
  5. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  6. Brom S., García de los Santos A., Stepkowsky T., Flores M., Dávila G., Romero D., Palacios R. Different plasmids of Rhizobium leguminosarum bv. phaseoli are required for optimal symbiotic performance. J Bacteriol. 1992 Aug;174(16):5183–5189. doi: 10.1128/jb.174.16.5183-5189.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Buchanan-Wollaston V., Cannon M. C., Beynon J. L., Cannon F. C. Role of the nifA gene product in the regulation of nif expression in Klebsiella pneumoniae. Nature. 1981 Dec 24;294(5843):776–778. doi: 10.1038/294776a0. [DOI] [PubMed] [Google Scholar]
  8. David M., Domergue O., Pognonec P., Kahn D. Transcription patterns of Rhizobium meliloti symbiotic plasmid pSym: identification of nifA-independent fix genes. J Bacteriol. 1987 May;169(5):2239–2244. doi: 10.1128/jb.169.5.2239-2244.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dover G. A. Evolution of genetic redundancy for advanced players. Curr Opin Genet Dev. 1993 Dec;3(6):902–910. doi: 10.1016/0959-437x(93)90012-e. [DOI] [PubMed] [Google Scholar]
  10. Eckhardt T. A rapid method for the identification of plasmid desoxyribonucleic acid in bacteria. Plasmid. 1978 Sep;1(4):584–588. doi: 10.1016/0147-619x(78)90016-1. [DOI] [PubMed] [Google Scholar]
  11. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  12. Figurski D. H., Helinski D. R. Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1648–1652. doi: 10.1073/pnas.76.4.1648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fischer H. M., Babst M., Kaspar T., Acuña G., Arigoni F., Hennecke H. One member of a gro-ESL-like chaperonin multigene family in Bradyrhizobium japonicum is co-regulated with symbiotic nitrogen fixation genes. EMBO J. 1993 Jul;12(7):2901–2912. doi: 10.1002/j.1460-2075.1993.tb05952.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Flores M., González V., Brom S., Martínez E., Piñero D., Romero D., Dávila G., Palacios R. Reiterated DNA sequences in Rhizobium and Agrobacterium spp. J Bacteriol. 1987 Dec;169(12):5782–5788. doi: 10.1128/jb.169.12.5782-5788.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Freiberg C., Fellay R., Bairoch A., Broughton W. J., Rosenthal A., Perret X. Molecular basis of symbiosis between Rhizobium and legumes. Nature. 1997 May 22;387(6631):394–401. doi: 10.1038/387394a0. [DOI] [PubMed] [Google Scholar]
  16. Gangloff S., Zou H., Rothstein R. Gene conversion plays the major role in controlling the stability of large tandem repeats in yeast. EMBO J. 1996 Apr 1;15(7):1715–1725. [PMC free article] [PubMed] [Google Scholar]
  17. Girard M. L., Flores M., Brom S., Romero D., Palacios R., Dávila G. Structural complexity of the symbiotic plasmid of Rhizobium leguminosarum bv. phaseoli. J Bacteriol. 1991 Apr;173(8):2411–2419. doi: 10.1128/jb.173.8.2411-2419.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Harvey S., Hill C. W. Exchange of spacer regions between rRNA operons in Escherichia coli. Genetics. 1990 Aug;125(4):683–690. doi: 10.1093/genetics/125.4.683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jackson J. A., Fink G. R. Gene conversion between duplicated genetic elements in yeast. Nature. 1981 Jul 23;292(5821):306–311. doi: 10.1038/292306a0. [DOI] [PubMed] [Google Scholar]
  20. Jones J. D., Gutterson N. An efficient mobilizable cosmid vector, pRK7813, and its use in a rapid method for marker exchange in Pseudomonas fluorescens strain HV37a. Gene. 1987;61(3):299–306. doi: 10.1016/0378-1119(87)90193-4. [DOI] [PubMed] [Google Scholar]
  21. Klein H. L., Petes T. D. Intrachromosomal gene conversion in yeast. Nature. 1981 Jan 15;289(5794):144–148. doi: 10.1038/289144a0. [DOI] [PubMed] [Google Scholar]
  22. Kobayashi I. Mechanisms for gene conversion and homologous recombination: the double-strand break repair model and the successive half crossing-over model. Adv Biophys. 1992;28:81–133. doi: 10.1016/0065-227x(92)90023-k. [DOI] [PubMed] [Google Scholar]
  23. Kündig C., Hennecke H., Göttfert M. Correlated physical and genetic map of the Bradyrhizobium japonicum 110 genome. J Bacteriol. 1993 Feb;175(3):613–622. doi: 10.1128/jb.175.3.613-622.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mahan M. J., Roth J. R. Role of recBC function in formation of chromosomal rearrangements: a two-step model for recombination. Genetics. 1989 Mar;121(3):433–443. doi: 10.1093/genetics/121.3.433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Margolin W., Long S. R. Rhizobium meliloti contains a novel second homolog of the cell division gene ftsZ. J Bacteriol. 1994 Apr;176(7):2033–2043. doi: 10.1128/jb.176.7.2033-2043.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Marsh J. L., Erfle M., Wykes E. J. The pIC plasmid and phage vectors with versatile cloning sites for recombinant selection by insertional inactivation. Gene. 1984 Dec;32(3):481–485. doi: 10.1016/0378-1119(84)90022-2. [DOI] [PubMed] [Google Scholar]
  27. Martínez-Salazar J. M., Romero D., Girard M. L., Dávila G. Molecular cloning and characterization of the recA gene of Rhizobium phaseoli and construction of recA mutants. J Bacteriol. 1991 May;173(10):3035–3040. doi: 10.1128/jb.173.10.3035-3040.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Noel K. D., Sanchez A., Fernandez L., Leemans J., Cevallos M. A. Rhizobium phaseoli symbiotic mutants with transposon Tn5 insertions. J Bacteriol. 1984 Apr;158(1):148–155. doi: 10.1128/jb.158.1.148-155.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Okazaki N., Matsuo S., Saito K., Tominaga A., Enomoto M. Conversion of the Salmonella phase 1 flagellin gene fliC to the phase 2 gene fljB on the Escherichia coli K-12 chromosome. J Bacteriol. 1993 Feb;175(3):758–766. doi: 10.1128/jb.175.3.758-766.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Orosz L., Páy A., Dallmann G. Heterozygosis of phage 16-3 of Rhizobium meliloti: moderate level of mismatch repair or gene conversion. Mol Gen Genet. 1980;179(1):163–167. doi: 10.1007/BF00268459. [DOI] [PubMed] [Google Scholar]
  31. Pardo M. A., Lagunez J., Miranda J., Martínez E. Nodulating ability of Rhizobium tropici is conditioned by a plasmid-encoded citrate synthase. Mol Microbiol. 1994 Jan;11(2):315–321. doi: 10.1111/j.1365-2958.1994.tb00311.x. [DOI] [PubMed] [Google Scholar]
  32. Prentki P., Krisch H. M. In vitro insertional mutagenesis with a selectable DNA fragment. Gene. 1984 Sep;29(3):303–313. doi: 10.1016/0378-1119(84)90059-3. [DOI] [PubMed] [Google Scholar]
  33. Quinto C., De La Vega H., Flores M., Leemans J., Cevallos M. A., Pardo M. A., Azpiroz R., De Lourdes Girard M., Calva E., Palacios R. Nitrogenase reductase: A functional multigene family in Rhizobium phaseoli. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1170–1174. doi: 10.1073/pnas.82.4.1170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Romero D., Brom S., Martínez-Salazar J., Girard M. L., Palacios R., Dávila G. Amplification and deletion of a nod-nif region in the symbiotic plasmid of Rhizobium phaseoli. J Bacteriol. 1991 Apr;173(8):2435–2441. doi: 10.1128/jb.173.8.2435-2441.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Romero D., Martínez-Salazar J., Girard L., Brom S., Dávilla G., Palacios R., Flores M., Rodríguez C. Discrete amplifiable regions (amplicons) in the symbiotic plasmid of Rhizobium etli CFN42. J Bacteriol. 1995 Feb;177(4):973–980. doi: 10.1128/jb.177.4.973-980.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schlüter A., Patschkowski T., Quandt J., Selinger L. B., Weidner S., Krämer M., Zhou L., Hynes M. F., Priefer U. B. Functional and regulatory analysis of the two copies of the fixNOQP operon of Rhizobium leguminosarum strain VF39. Mol Plant Microbe Interact. 1997 Jul;10(5):605–616. doi: 10.1094/MPMI.1997.10.5.605. [DOI] [PubMed] [Google Scholar]
  37. Segall A. M., Roth J. R. Approaches to half-tetrad analysis in bacteria: recombination between repeated, inverse-order chromosomal sequences. Genetics. 1994 Jan;136(1):27–39. doi: 10.1093/genetics/136.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Segovia L., Young J. P., Martínez-Romero E. Reclassification of American Rhizobium leguminosarum biovar phaseoli type I strains as Rhizobium etli sp. nov. Int J Syst Bacteriol. 1993 Apr;43(2):374–377. doi: 10.1099/00207713-43-2-374. [DOI] [PubMed] [Google Scholar]
  39. Simon R. High frequency mobilization of gram-negative bacterial replicons by the in vitro constructed Tn5-Mob transposon. Mol Gen Genet. 1984;196(3):413–420. doi: 10.1007/BF00436188. [DOI] [PubMed] [Google Scholar]
  40. Szostak J. W., Wu R. Unequal crossing over in the ribosomal DNA of Saccharomyces cerevisiae. Nature. 1980 Apr 3;284(5755):426–430. doi: 10.1038/284426a0. [DOI] [PubMed] [Google Scholar]
  41. Valderrama B., Dávalos A., Girard L., Morett E., Mora J. Regulatory proteins and cis-acting elements involved in the transcriptional control of Rhizobium etli reiterated nifH genes. J Bacteriol. 1996 Jun;178(11):3119–3126. doi: 10.1128/jb.178.11.3119-3126.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Williams S. M., Strobeck C. Sister chromatid exchange and the evolution of rDNA spacer length. J Theor Biol. 1985 Oct 21;116(4):625–636. doi: 10.1016/s0022-5193(85)80092-8. [DOI] [PubMed] [Google Scholar]
  43. Yamamoto K., Takahashi N., Fujitani Y., Yoshikura H., Kobayashi I. Orientation dependence in homologous recombination. Genetics. 1996 May;143(1):27–36. doi: 10.1093/genetics/143.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Yamamoto K., Yoshikura H., Takahashi N., Kobayashi I. Apparent gene conversion in an Escherichia coli rec+ strain is explained by multiple rounds of reciprocal crossing-over. Mol Gen Genet. 1988 Jun;212(3):393–404. doi: 10.1007/BF00330842. [DOI] [PubMed] [Google Scholar]

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