Comparative Proteomic Analysis Revealing ActJ-Regulated Proteins in Sinorhizobium meliloti

Abstract

To adapt to different environmental conditions,Sinorhizobium meliloti relies on finely tuned regulatory networks,most of which are unexplored to date. We recently demonstratedthat deletion of the two-component system ActJK renders an acidvulnerable phenotype in S. meliloti and negatively impactsbacteroid development and nodule occupancy as well. To fullyunderstand the role of ActJ in acid tolerance, S. meliloti wild-typeand S. meliloti ΔactJ proteomes were compared in the presence orabsence of acid stress by nanoflow ultrahigh-performance liquidchromatography coupled to mass spectrometry. The analysisdemonstrated that proteins involved in the synthesis ofexopolysaccharides (EPSs) were notably enriched in ΔactJ cellsin acid pH. Total EPS quantification further revealed that although EPS production was augmented at pH 5.6 in both the ΔactJ andthe parental strain, the lack of ActJ significantly enhanced this difference. Moreover, several efflux pumps were found to bedownregulated in the ΔactJ strain. Promoter fusion assays suggested that ActJ positively modulated its own expression in an acidmedium but not at under neutral conditions. The results presented here identify several ActJ-regulated genes in S. meliloti,highlighting key components associated with ActJK regulation that will contribute to a better understanding of rhizobia adaptation toacid stress.