Regulation of cysteine residues in LsrB proteins from Sinorhizobium meliloti under free‐living and symbiotic oxidative stress

Abstract

The development of legume nitrogen-fixing nodules is regulated by reactive oxygen species (ROS) produced by symbionts. Several regulators from Rhizobium are involved in ROS sensing. In a previous study, we found that Sinorhizobium meliloti LsrB regulates lipopolysaccharide production and is associated with H O accumulation in alfalfa (Medicago sativa) nodules. However, its underlying regulatory mechanism remains unclear. Here, we report that the cysteine residues in LsrB are required for adaptation to oxidative stress, gene expression, alfalfa nodulation and nitrogen fixation. Moreover, LsrB directly activated the transcription of lrp3 and gshA (encoding γ-glutamylcysteine synthetase, responsible for glutathione synthesis) and this regulation required the cysteine (Cys) residues in the LsrB substrate-binding domain. The Cys residues could sense oxidative stress via the formation of intermolecular disulfide bonds, generating LsrB dimers and LsrB-DNA complexes. Among the Cys residues, C238 is a positive regulatory site for the induction of downstream genes, whereas C146 and C275 play negative roles in the process. The lsrB mutants with Cys-to-Ser substitutions displayed altered phenotypes in respect to their adaptation to oxidative stress, nodulation and nitrogen fixation-related plant growth. Our findings demonstrate that S. meliloti LsrB modulates alfalfa nodule development by directly regulating downstream gene expression via a post-translational strategy.

A central role for the transcriptional regulator VtlR in small RNA-mediated gene regulation in Agrobacterium tumefaciens

“…Regarding regulatory roles, S. meliloti LsrB is involved in the regulation of genes required for synthesizing glutathione and lipopolysaccharide 19 , 20 . Recently, S. meliloti LsrB was shown to have a similar sensing mechanism as OxyR, where the formation of intermolecular disulfide bonds in LsrB is involved in adaptation to oxidative stress, regulation of gene expression, proper alfalfa nodulation, and effective nitrogen fixation 21 .…”

Involvement of Glutaredoxin and Thioredoxin Systems in the Nitrogen-Fixing Symbiosis between Legumes and Rhizobia

“…LsrB belongs to the LysR family of bacterial transcriptional regulators including the oxidative stress regulator OxyR [81]. An lsrB deletion mutant has a reduced pool of GSH, and LsrB inactivation accordingly results in the decreased expression of genes involved in GSH metabolism ( gshA , gshB , gor ) both in free-living and in planta [82,83]. The regulator was shown to directly activate the expression of gshA , and to respond to cellular redox changes via the three reactive cysteines in the substrate-binding domain [83].…”

“…An lsrB deletion mutant has a reduced pool of GSH, and LsrB inactivation accordingly results in the decreased expression of genes involved in GSH metabolism ( gshA , gshB , gor ) both in free-living and in planta [82,83]. The regulator was shown to directly activate the expression of gshA , and to respond to cellular redox changes via the three reactive cysteines in the substrate-binding domain [83]. LsrB also positively regulates the expression of genes involved in lipopolysaccharide biosynthesis [84].…”

“…LsrB also positively regulates the expression of genes involved in lipopolysaccharide biosynthesis [84]. Nodules induced by mutants defective in LsrB undergo premature senescence coupled to impaired bacteroid differentiation and ROS accumulation, which could be partly due to GSH deficiency [83]. Several genes of the Trx/Grx systems belong to the RpoH1 regulon.…”

Molecular Weapons Contribute to Intracellular Rhizobia Accommodation Within Legume Host Cell

“…Besides, nodules induced by a Sinorhizobium meliloti deletion mutant of lsrB , which encodes a LysR transcription factor acting as a ROS regulator, showed premature senescence with impaired bacteroid differentiation (Luo et al, 2005; Tang et al, 2013). LsrB was found to induce the expression of the lrp3 - lpsCDE operon involved in lipopolysaccharide biosynthesis required for infection or bacteroid survival in host cells ( Figure 1 )(Tang et al, 2014) and that of γ-glutamylcysteine synthetase, involved in glutathione synthesis (Tang et al, 2017).…”