The regulatory status of the fixL- and fixJ-like genes in Bradyrhizobium japonicum may be different from that in Rhizobium meliloti

Abstract

The cloning, sequencing and mutational analysis of the Bradyrhizobium japonicum symbiotic nitrogen fixation genes fixL and fixJ are reported here. The two genes were adjacent and probably formed an operon, fixLJ. The predicted FixL and FixJ proteins, members of the two-component sensor/regulator family, were homologous over almost their entire lengths to the corresponding Rhizobium meliloti proteins (approx. 50% identity). Downstream of the B. japonicum fixJ gene was found an open reading frame with 138 codons (ORF138) whose product shared 36% homology with the N-terminal part of FixJ. Deletion and insertion mutations within fixL and fixJ led to a loss of approximately 90% wild-type symbiotic nitrogen fixation (Fix) activity, whereas an ORF138 mutant was Fix+. In fixL, fixJ and ORF138 mutant backgrounds, the aerobic expression of the fixR-nifA operon was not affected. NifA itself did not regulate the expression of the fixJ gene. Thus, the B. japonicum FixL and FixJ proteins were neither involved in the regulation of aerobic nifA gene expression nor in the anaerobic NifA-dependent autoregulation of the fixRnifA operon, rather they appeared to control symbiotically important genes other than those whose expression was dependent on the NifA protein. The fixL and fixJ mutant strains were unable to grow anaerobically with nitrate as the terminal electron acceptor. Therefore, some of the FixJ-dependent genes in B. japonicum may be concerned with anaerobic respiration.

Characterization of a fixLJ-regulated Bradyrhizobium japonicum gene sharing similarity with the Escherichia coli fnr and Rhizobium meliloti fixK genes

“…Expression of B. japonicum fixK is controlled by the FixlJ system. Recently, the fixLJ genes were identified in B. japonicum (2). To test whether expression of the fixK-like gene of B. japonicum depends on the oxygen conditions and is regulated by the FixLJ system, a translational fixK'-'lacZ fusion was constructed (see Materials and Methods).…”

Genetic and functional characterization of the gene cluster directing the biosynthesis of putisolvin I and II in Pseudomonas putida strain PCL1445

“…The HTH motif has been observed in many regulatory proteins (Pabo & Sauer, 1992), which are divided into more than 10 groups, including the LuxR, AraC and MarR families. PsoR appears to be most closely related to members of the LuxR regulatory family, such as DctR (Hamblin et al, 1993) and FixJ (Anthamatten & Hennecke, 1991). An approximately 60 aa region of the C terminus containing the four helices and their turns, which is called a three-element fingerprint, provides the signature for the HTH motif of the LuxR family of bacterial regulatory proteins.…”

“…Despite this homology, PsoR lacks five highly conserved residues at the N terminus characteristic of the LuxR subfamily, which is composed of autoinducerbinding regulators activated by homoserine lactones (Fuqua et al, 1996). The second major subfamily of transcriptional regulators is composed of the response regulators of twocomponent signal transduction systems, such as FixJ (Anthamatten & Hennecke, 1991) and DctR (Hamblin et al, 1993). Three highly conserved residues (Arg, Glu, Lys) characteristic of the response regulators were found in the PsoR sequence, suggesting that PsoR may be closely related to this subfamily of regulators (Fig.…”

“…Database searches with BLASTP demonstrated that approximately 60 aa in the C-terminal region of PsoR shared significant similarity with similar regions of known regulatory proteins. The C terminus of PsoR exhibited 35 % identity to GerE (Cutting & Mandelstam, 1986), 34 % identity to DctR (Hamblin et al, 1993) and 32 % identity to FixJ (Anthamatten & Hennecke, 1991) (Fig. 5a).…”

Oxygen Signal Transduction

“…Four FixL/FixJ couples have been reported so far, those of S. meliloti, Bradyrhizobium japonicum, Rhizobium leguminosarum, and Azorhizobium caulinodans (10,(16)(17)(18). These Rhizobia are, respectively, the symbiotic partners of alfalfa, soybean, peas, and Sesbania rostrata.…”