Abstract

Quorum sensing (QS) is a process in which bacteria use diverse signaling molecules to monitor their population density and regulate population-wide expression of genes involved in several key bacterial processes such as virulence, biofilm formation, and antibiotic resistance. Gram-positive bacteria typically use oligopeptides as intercellular signaling molecules. The secreted oligopeptides modulate gene expression by either activating the sensor kinase of a two-component system on the bacterial surface or by interacting with cognate transcription regulator in the bacterial cytosol. The gram-positive bacteria Group A Streptococcus (GAS) is a major human pathogen responsible for over 700 million infections annually worldwide. GAS produces a wide spectrum of virulence factors that play crucial roles in disease pathogenesis. Among the many toxins produced by GAS, Streptococcal pyrogenic exotoxin B (SpeB) is one of the well-studied virulence factor. SpeB is a secreted cysteine protease that is produced abundantly during infection and is critical for GAS pathogenesis. Although SpeB is extensively studied for a century, the precise regulatory events that govern speB gene expression are not fully understood. In this study, we have discovered that GAS employs a previously unknown peptide-mediated quorum sensing pathway to control speB expression during high bacterial population density. GAS genome encodes a novel class of leaderless peptide signal, SpeB-Inducing Peptide (SIP). SIP lacks several characteristic features that are hallmarks of bacterial peptide signals. Contrary to all the characterized bacterial peptide signals, SIP is produced in its mature form and lacks amino acid sequences in the amino terminus required for secretion. Nevertheless, SIP functions as an effective intercellular signal. SIP is secreted and reinternalized into GAS cytosol where it interacts with its cognate regulator, Regulator of proteinase B (RopB). SIP binding to RopB induces allosteric changes in the regulator, which leads to high affinity RopB-DNA interactions, RopB oligomerization and activation of speB gene expression. Importantly, we demonstrate that the SIP signaling pathway is active in vivo and contributes significantly to GAS virulence in multiple mouse models of GAS infection. We also show that the SIP signaling occurs during GAS growth ex vivo in human saliva and blood and SIP-mediated speB expression is crucial for GAS survival in both saliva and blood. Together, our discoveries in this study identify a novel bacterial signaling pathway and suggest new therapeutic strategies for future translation studies.