Abstract
We have previously demonstrated that the Rhizobium meliloti large plasmid pRmeGR4b carries the gene locus nodule formation efficiency (nfe) which is responsible for nodulation efficiency and competitive ability of strain GR4 on alfalfa roots. In this study we report that expression of nfe-lacZ fusions in Escherichia coli is activated in the presence of the cloned nifA gene of R. meliloti. This activation was found to be oxygen sensitive and to require the E. coli ntrA gene product. In contrast to the R. meliloti nifA, the cloned nifA gene of Klebsiella pneumoniae was able to activate expression of nfe in acrobically grown cells of both E. coli and R. meliloti. Hybridization experiments did not show homology to nfe in four R. meliloti wild-type strains tested. These strains were uncompetitive when coinoculated with a GR4 derivative carrying plasmid pRmeGR4b, but were competitive when coinoculated with a GR4 derivative carrying a single transposon mutation into the nfe region. When nfe DNA was introduced into the four wild-type strains, a significant increase in the competitive ability of two of them was observed, as deduced from their respective percentages of alfalfa root nodule occupancy in twostrains coinoculation experiments.
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References
Batut J, Terzaghi B, Ghérardi M, Huguet M, Terzaghi E, Garnerone AM, Boistard P, Huguet T (1985) Localization of a symbiotic fix region on R. meliloti pSym megaplasmid more than 200 kilobases from the nod-nif region. Mol Gen Genet 199:232–239
Better M, Lewis B, Corbin D, Ditta G, Helinski DR (1985) Deletion analysis of R. meliloti symbiotic promoters. EMBO J 4:2419–2424
Beynon JL, Williams MK, Cannon FC (1988) Expression and functional analysis of the Rhizobium meliloti nif A gene. EMBO J 7:7–14
Boyer HW, Roulland-Dussoix D (1969) A complementation analysis of the restriction and modification of DNA in E. coli. J Mol Biol 41:459–465
Cannon FC, Dixon RA, Postgate JR, Primrose SB (1974) Chromosomal integration of Klebsiella pneumoniae nitrogen fixation genes in Escherichia coli. J Gen Microbiol 80:227–239
David M, Daveran M, Batut J, Dedieu A, Domergue O, Ghai J, Hertig C, Boistard P, Khan D (1988) Cascade regulation of nif gene expression in Rhizobium meliloti. Cell 54:671–683
Ditta G, Stanfield S, Corbin D, Helinski DR (1980) Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium meliloti. Proc Natl Acad Sci USA 77:7347–7351
Ditta G, Virts E, Palomares AJ, Kim CH (1987) The nifA gene of Rhizobium meliloti is oxygen regulated. J Bacteriol 169:3217–3223
Dowling DN, Broughton WJ (1986) Competition for nodulation of legumes. Annu Rev Microbiol 40:131–157
Eckhardt TH (1978) A rapid method for the identification of plasmid deoxyribonucleic acid in bacteria. Plasmid 1:584–588
Figurski DH, Helinski DR (1979) Replication of an origin-containing derivative of plasmid RK2 dependent on a function provided in trans. Proc Natl Acad Sci USA 76:1648–1652
Fischer HM, Alvarez-Morales A, Hennecke H (1986) The pleiotropic nature of symbiotic regulatory mutants: Bradyrhizobium japonicum nif A gene is involved in control of nif gene expression and formation of determinate symbiosis. EMBO J 5:1165–1173
Gussin GN, Ronson CW, Ausubel FM (1986) Regulation of nitrogen fixation genes. Annu Rev Genet 20:567–591
Hill S, Kennedy C, Kavanagh E, Goldberg RB, Hanau R (1981) Nitrogen fixation gene (nifL) involved in oxygen regulation of nitrogenase synthesis in Klebsiella pneumoniae. Nature 200:424–426
Hirsch AM, Smith CA (1987) Effects of Rhizobium meliloti nif and fix mutants on alfalfa root nodule development. J Bacteriol 169:1137–1146
Kennedy C, Robson RL (1983) Activation of nif gene expression in Azotobacter by the nif A gene product of Klebsiella pneumoniae. Nature 301:626–628
Landgridge J, Landgridge P, Bergquist PL (1980) Extraction of nucleic acids from agarose gels. Anal Biochem 103:264–271
Long SR, Buikema WJ, Ausubel FM (1982) Cloning of Rhizobium meliloti nodulation genes by direct complementation of Nod- mutants. Nature 298:485–488
MacNeil T, Roberts GP, MacNeil D, Tyler B (1982) The products of glnL and glnG are bifunctional regulatory proteins. Mol Gen Genet 188:325–333
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning; a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Mcloughlin TJ, Merlo AO, Satola SW, Johansen S (1987) Isolation of competition-defective mutants of Rhizobium fredii. J Bacteriol 169:410–413
Merrick M, Hill S, Hennecke H, Hahn M, Dixon R, Kennedy C (1982) Repressor properties of the nifL gene product in Klebsiella pneumoniae. Mol Gen Genet 185:75–81
Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Olivares J, Casadesús J, Bedmar EJ (1980) Method for testing degree of infectivity of Rhizobium meliloti strains. Appl Environ Microbiol 56:389–393
Oliveira LA, Graham PH (1990) Speed of nodulation and competitive ability among strins of Rhizobium leguminosarum bv phaseoli. Arch Microbiol 153:311–315
Rigby PW, Dieckman M, Rhodes C, Berg P (1977) Labelling deoxyribonucleic acid to high specific activity in vitro nick translation with DNA polymerase. J Mol Biol 113:237–251
Ronson CW, Nixon BT, Albright L, Ausubel FM (1987) Rhizobium meliloti ntrA (rpoN) gene is required for diverse metabolic functions. J Bacteriol 169:2424–2431
Ruvkun GB, Sundaresan V, Ausubel FM (1982) Directed transposon Tn5 mutagenesis and complementation analysis of Rhizobium meliloti symbiotic nitrogen fixation genes. Cell 29:551–559
Sanjuan J, Olivares J (1989) Implication of nif A in the regulation of genes located on a Rhizobium meliloti cryptic plasmid that affect nodulation efficiency. J Bacteriol 171:4154–4161
Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517
Szeto WW, Zimmerman J, Sundaresan V, Ausubel FM (1984) A Rhizobium meliloti symbiotic regulatory gene. Cell 36:1035–1043
Toro N, Olivares J (1986) Characterization of a large plasmid of Rhizobium meliloti involved in enhancing nodulation. Mol Gen Genet 202:331–335
Weber G, Reiländer H, Pühler A (1985) Mapping and expression of a regulatory nitrogen fixation gene (fixD) of Rhizobium meliloti. EMBO J 4:2751–2756
Zdor RE, Pueppke SG (1988) Early infection and competition for nodulation of soybean by Bradyrhizobium japonicum 123 and 138. Appl Environ Microbiol 54:1196–2002
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Sanjuan, J., Olivares, J. NifA-NtrA regulatory system activates transcription of nfe, a gene locus involved in nodulation competitiveness of Rhizobium meliloti . Arch. Microbiol. 155, 543–548 (1991). https://doi.org/10.1007/BF00245347
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DOI: https://doi.org/10.1007/BF00245347