Functional Characterization of RsRsgA for Ribosome Biosynthesis and Expression of the Type III Secretion System in Ralstonia solanacearum

Abstract

RsgA plays an important role in maturation of 30S subunit in many bacteria that assists in the release of RbfA from the 30S subunit during a late stage of ribosome biosynthesis. Here, we genetically characterized functional roles of RsgA in Ralstonia solanacearum, hereafter designated RsRsgA. Deletion of R. solanacearum rsgA or rbfA resulted in distinct deficiency of 16S ribosomal RNA, significantly slowed growth in broth medium, and diminished growth in nutrient-limited medium, which are similar as phenotypes of rsgA mutants and rbfA mutants of Escherichia coli and other bacteria. Our gene-expression studies revealed that RsRsgA is important for expression of genes encoding the type III secretion system (T3SS) (a pathogenicity determinant of R. solanacearum) both in vitro and in planta. Compared with the wild-type R. solanacearum strain, proliferation of the rsgA and rbfA mutants in tobacco leaves was significantly impaired, while they failed to migrate into tobacco xylem vessels from infiltrated leaves, and hence, these two mutants failed to cause any bacterial wilt disease in tobacco plants. It was further revealed that rsgA expression was highly enhanced under nutrient-limited conditions compared with that in broth medium and RsRsgA affects T3SS expression through the PrhN-PrhG-HrpB pathway. Moreover, expression of a subset of type III effectors was substantially impaired in the rsgA mutant, some of which are responsible for R. solanacearum GMI1000 elicitation of a hypersensitive response (HR) in tobacco leaves, while RsRsgA is not required for HR elicitation of GMI1000 in tobacco leaves. All these results provide novel insights into understanding various biological functions of RsgA proteins and complex regulation on the T3SS in R. solanacearum.