The rkpU gene of Sinorhizobium fredii HH103 is required for bacterial K-antigen polysaccharide production and for efficient nodulation with soybean but not with cowpea

Abstract

In this work, the role of the rkpU and rkpJ genes in the production of the K-antigen polysaccharides (KPS) and in the symbiotic capacity of Sinorhizobium fredii HH103, a broad host-range rhizobial strain able to nodulate soybean and many other legumes, was studied. The rkpJ-and rkpU-encoded products are orthologous to Escherichia coli proteins involved in capsule export. S. fredii HH103 mutant derivatives were contructed in both genes. To our knowledge, this is the first time that the role of rkpU in KPS production has been studied in rhizobia. Both rkpJ and rkpU mutants were unable to produce KPS. The rkpU derivative also showed alterations in its lipopolysaccharide (LPS). Neither KPS production nor rkpJ and rkpU expression was affected by the presence of the flavonoid genistein. Soybean (Glycine max) plants inoculated with the S. fredii HH103 rkpU and rkpJ mutants showed reduced nodulation and clear symptoms of nitrogen starvation. However, neither the rkpJ nor the rkpU mutants were significantly impaired in their symbiotic interaction with cowpea (Vigna unguiculata). Thus, we demonstrate for the first time to our knowledge the involvement of the rkpU gene in rhizobial KPS production and also show that the symbiotic relevance of the S. fredii HH103 KPS depends on the specific bacterium-legume interaction.

Sinorhizobium fredii HH103 does not strictly require KPS and/or EPS to nodulate Glycyrrhiza uralensis, an indeterminate nodule-forming legume

“…At the present, four S. fredii HH103 KPS mutants (in rkpH, rkpG, rkpJ, and rkpU) have been investigated for their capacity to produce KPS and for their symbiotic properties with Glycine max (Hidalgo et al 2010;Parada et al 2006). In this work, two other S. fredii HH103 KPS mutants (in rkpA and rkpI) were constructed and their characteristics investigated.…”

“…In G. max, a determinate noduleforming legume, only KPS is symbiotically relevant (Parada et al 2006). In Vigna unguiculata, another determinate nodule-forming legume, neither KPS nor EPS are required for a successful symbiotic interaction (Hidalgo et al 2010).…”

“…For detection of KPS, crude extracts from S. fredii mutants SVQ536 (rkpA), SVQ536 carrying cosmid pMUS766, SVQ538 (rkpI), and SVQ538 carrying cosmid pMUS759 were obtained and analyzed by NMR as previously described (Crespo-Rivas et al 2009;Hidalgo et al 2010). For KPS visualization, samples were prepared as follows: bacterial cultures were grown on liquid TY medium for 48 h. Bacterial cells were washed 3 times in 0.9% NaCl and pelleted by centrifugation.…”

“…Four S. fredii genes belonging to the rkp-1 region have been mutated (rkpG, rkpH, rkpJ, and rkpU) and the symbiotic properties of their mutant derivatives reported (Hidalgo et al 2010;Parada et al 2006). All this S. fredii HH103 rkp mutants fail to produce KPS and their symbiotic capacity with Glycine max (soybean) is severely reduced.…”

“…The rkpJ gene is homologous to the Escherichia coli kpsS gene, which codes for a putative capsular polysaccharide transport protein. The kpsF3 codes for a putative capsule expression protein (Hidalgo et al 2010;Kiss et al 1997;Parada et al 2006;Petrovics et al 1993).…”