The Rhizobium leguminosarum bv. trifolii RosR: Transcriptional Regulator Involved in Exopolysaccharide Production

Abstract

The acidic exopolysaccharide is required for the establishment of symbiosis between the nitrogen-fixing bacterium Rhizobium leguminosarum bv. trifolii and clover. Here, we describe RosR protein from R. leguminosarum bv. trifolii 24.2, a homolog of transcriptional regulators belonging to the family of Ros/MucR proteins. R. leguminosarum bv. trifolii RosR possesses a characteristic Cys2His2 type zinc-finger motif in its C-terminal domain. Recombinant (His)6RosR binds to an RosR-box sequence located up-stream of rosR. Deletion analysis of the rosR upstream region resulted in identification of two -35 to -10 promoter sequences, two conserved inverted palindromic pentamers that resemble the cAMP-CRP binding site of Escherichia coli, inverted repeats identified as a RosR binding site, and other regulatory sequence motifs. When assayed in E. coli, a transcriptional fusion of the cAMP-CRP binding site containing the rosR upstream region and lacZ gene was moderately responsive to glucose. The sensitivity of the rosR promoter to glucose was not observed in E. coli deltacyaA. A rosR frame-shift mutant of R. leguminosarum bv. trifolii formed dry, wrinkled colonies and induced nodules on clover, but did not fix nitrogen. In the rosR mutant, transcription of pssA-lacZ fusion was decreased, indicating positive regulation of the pssA gene by RosR. Multiple copies of rosR in R. leguminosarum bv. trifolii 24.2 increased exopolysaccharide production.

Diverse Genetic Regulon of the Virulence-Associated Transcriptional Regulator MucR in Brucella abortus 2308

“…For example, a naturally mucoid colony of Agrobacterium tumefaciens loses its mucoid appearance when ros is mutated (5), and a mucR mutation in Sinorhizobium meliloti results in a similar loss of colony mucoidy (8). Importantly, these mucoidy-related phenotypes have been linked to the disregulation of genes encoding proteins involved in exopolysaccharide synthesis (11,12,13,14,15). The B. abortus mucR mutant does not appear to be any more or less mucoid than parental strain 2308 based on macroscopic observations, but it is interesting that MucR controls the expression of genes (bab1_1973 to bab1_1975) homologous to the exsB, -C, and -D genes of S. meliloti, which are involved in succinoglycan production (40).…”

“…While early studies genetically linked ros and mucR mutations to growth defects and differences in colony mucoidy, the mechanism of action of the Ros/MucR proteins was not known at the time; however, it has since been determined that Ros/MucR proteins are transcriptional repressors that regulate numerous genes, including those involved in polysaccharide synthesis, motility, and quorum sensing (11,12,13,14,15,16,17,18). Ros/MucRtype regulators are unusual in that they contain a Zn finger motif that is uncommon in prokaryotes, whereas transcriptional regulators with this type of motif are commonly found in eukaryotes (19).…”

The response of the Rhizobium leguminosarum bv. trifolii wild-type and exopolysaccharide-deficient mutants to oxidative stress

“…Previously, we have established that the transcriptional regulator RosR of R. leguminosarum bv. trifolii positively regulates the pssA expression (Janczarek and Skorupska 2007;Janczarek and Urbanik-Sypniewska 2013). This gene encodes a protein, which besides its enzymatic activity is probably involved additionally in other cellular processes (Janczarek and Skorupska 2003).…”

“…pssA encodes a glucosyl-IP-transferase involved in the first step of EPS synthesis and a mutation in this gene results in a total lack of production of this polymer. rosR encodes a transcriptional regulator involved in positive regulation of this process and its mutation substantially (3-fold) decreases the amount of produced EPS (Janczarek and Skorupska 2007;Janczarek et al 2009a). Both mutants induce formation of small nodules on clover plants that are unable to fix nitrogen.…”

Do Ca2+-chelating polysaccharides reduce calcium ion release from gypsum-based biomaterials?

“…All other chemicals of analytical grade were supplied by POCH, Gliwice (Poland). EPS CE3 was isolated from supernatants of 72 h R. etli CE3 cultures, as was described previously [26]. The content of Ca ions in both polysaccharides was assayed using a commercial kit (Calcium CPC, Biomaxima, Poland) prior to the experiments; both preparations were found to be calcium-free.…”