The pleiotropic nature of symbiotic regulatory mutants: Bradyrhizobium japonicum nifA gene is involved in control of nif gene expression and formation of determinate symbiosis

Abstract

In the slow‐growing soybean symbiont, Bradyrhizobium japonicum (strain 110), a nifA‐like regulatory gene was located immediately upstream of the previously mapped fixA gene. By interspecies hybridization and partial DNA sequencing the gene was found to be homologous to nifA from Klebsiella pneumoniae and Rhizobium meliloti, and to a lesser extent, also to ntrC from K. pneumoniae. The B. japonicum nifA gene product was shown to activate B. japonicum and K. pneumoniae nif promoters (using nif::lacZ translational fusions) both in Escherichia coli and B. japonicum backgrounds. In the heterologous E. coli system activation was shown to be dependent on the ntrA gene product. Site‐directed insertion and deletion/replacement mutagenesis revealed that nifA is probably the promoter‐distal cistron within an operon. NifA‐ mutants were Fix‐ and pleiotropic: (i) they were defective in the synthesis of several proteins including the nifH gene product (nitrogenase Fe protein); the same proteins had been known to be repressed under aerobic growth of B. japonicum but derepressed at low O2 tension; (ii) the mutants had an altered nodulation phenotype inducing numerous, small, widely distributed soybean nodules in which the bacteroids were subject to severe degradation. These results show that nifA not only controls nitrogenase genes but also one or more genes involved in the establishment of a determinate, nitrogen‐fixing root nodule symbiosis.

Critical spacing between two essential cysteine residues in the interdomain linker of the Bradyrhizobium japonicum NifA protein

“…NifA activity was assayed in an E. coli in vivo system by measuring NifA-dependent activation of a translational nifD '-'lacZ fusion located on plasmid pRJ 1025 [14,15]. The test cultures were grown microaerobically in supplemented NFDM medium as described [3], and ~-galactosidase activity was determined according to Miller [16].…”

Sinorhizobium meliloti nifA mutant induces different gene expression profile from wild type in Alfalfa nodules

“…The nifA gene exerts its pleiotropic nature in controlling further nodulation by eliciting host plant defense reactions at early stages in development [3]. In addition to the nif/fix genes, four nodulation-specific genes (nodH, nodL, nolF and noeB) were down-regulated in 30-day nodules induced by the S. meliloti nifA mutant (unpublished data).…”

“…Previous studies inferred that nifA mutants not only failed to fix nitrogen but also elicited numerous small nodules whose necrotic interior was reminiscent of a hypersensitive response characteristic of non-compatible host-pathogen interactions [3]. The nodules induced by the S. meliloti nifA mutant differed from those induced by wild type in that the fixation zone was less extensive and many of the cells 819 npg interspersed within the fixation zone contained senescent bacteroids.…”

“…Nitrogen fixation in legumes takes place in highly specialized root nodules that result from the association between Rhizobia and legume plants [1][2][3]. The bacteria enter the nodule via an infection thread, and then differentiate into nitrogen-fixation bacteroids, which are capable of fixing atmospheric nitrogen into ammonia that provided to the plant.…”

Effects of Oxygen Levels on the Transcription of nif and gln Genes in Bradyrhizobium japonicum

“…The glnII gene promoter shares sequence homology in the -10 to -25 region with B. japonicum nifpromoters (Carlson et al, 1987) and with promoters from other bacteria that are controlled by nitrogen availability (Ntr control; Magasanik, 1982;Ausubel, 1984;Gussin et al, 1986). A gene, termed nifA, that is required for both microaerobic and symbiotic induction of nif and glnII gene expression in B. japonicum has been characterized (Fischer et al, 1986;Adams, 1986). The nifA gene product acts on a DNA sequence that is found upstream of nifgenes (Alvarez-Morales et al, 1986) but not upstream of the glnII gene (Carlson et al, 1987).…”