Rgg proteins associated with internalized small hydrophobic peptides: a new quorum‐sensing mechanism in streptococci

Abstract

SummaryWe identified a genetic context encoding a transcriptional regulator of the Rgg family and a small hydrophobic peptide (SHP) in nearly all streptococci and suggested that it may be involved in a new quorumsensing mechanism, with SHP playing the role of a pheromone. Here, we provide further support for this hypothesis by constructing a phylogenetic tree of the Rgg and Rgg-like proteins from Gram-positive bacteria and by studying the shp/rgg1358 locus of Streptococcus thermophilus LMD-9. We identified the shp1358 gene as a target of Rgg1358, and used it to confirm the existence of the steps of a quorumsensing mechanism including secretion, maturation and reimportation of the pheromone into the cell. We used surface plasmon resonance to demonstrate interaction between the pheromone and the regulatory protein and performed electrophoretic mobility shift assays to assess binding of the transcriptional regulator to the promoter regions of its target genes. The active form of the pheromone was identified by mass spectrometry. Our findings demonstrate that the shp/rgg1358 locus encodes two components of a novel quorum-sensing mechanism involving a transcriptional regulator of the Rgg family and a SHP pheromone that is detected and reimported into the cell by the Ami oligopeptide transporter.

Rgg protein structure–function and inhibition by cyclic peptide compounds

“…Cys45 forms the disulfide bond, and this cysteine is absolutely conserved among more than 120 Rgg2 and Rgg3 orthologs from 27 different species. In contrast, this cysteine is absent from the more than 400 proteins that comprise the other Rgg subfamilies, including the group II, group III, ComR, and RopB proteins (6).…”

“…Rgg proteins are widespread in Firmicute species, including but not limited to the Streptococcaceae, Lactobacillales, Listeriaceae, and Enterococcaceae (6). It also is common for organisms to express multiple paralogous Rgg proteins putatively serving nonredundant regulatory functions.…”

“…1A). Gram-positive cytoplasmic pheromone receptors include Bacillus response regulator aspartate phosphatases (Rap), neutral protease regulator (NprR), and phosphatidylinositol-specific phospholipase C gene regulator (PlcR), Enterococcus pheromone-responsive transcription factor (PrgX), and Streptococcus regulator gene of glucosyltransferase (Rgg) (as well as the homologous MutR and GadR) (4)(5)(6)(7)(8)(9)(10)(11). The Rap proteins are phosphatases and transcriptional antiactivators, whereas NprR, PlcR, and PrgX are DNA-binding transcription factors.…”

“…The 3D structure of Rgg proteins was unknown; however, based on their functional similarity to NprR, PrgX, and PlcR and their remote sequence similarity to RNPP family proteins, Rgg proteins preliminarily were included in this group (4,6,7,20).…”

Structural basis for the activation mechanism of the PlcR virulence regulator by the quorum-sensing signal peptide PapR

“…observed, pointing out their important role in adaptative and virulence processes (27)(28)(29)(30)(31)(32). This clearly identifies these regulators as major targets for the search of new molecules with applications in medical, food, and biotechnology areas.…”