Leaderless secreted peptide signaling molecule alters global gene expression and increases virulence of a human bacterial pathogen

Abstract

Successful pathogens use complex signaling mechanisms to monitor their environment and reprogram global gene expression during specific stages of infection. Group A (GAS) is a major human pathogen that causes significant disease burden worldwide. A secreted cysteine protease known as streptococcal pyrogenic exotoxin B (SpeB) is a key virulence factor that is produced abundantly during infection and is critical for GAS pathogenesis. Although identified nearly a century ago, the molecular basis for growth phase control of gene expression remains unknown. We have discovered that GAS uses a previously unknown peptide-mediated intercellular signaling system to control SpeB production, alter global gene expression, and enhance virulence. GAS produces an eight-amino acid leaderless peptide [SpeB-inducing peptide (SIP)] during high cell density and uses the secreted peptide for cell-to-cell signaling to induce population-wide expression. The SIP signaling pathway includes peptide secretion, reimportation into the cytosol, and interaction with the intracellular global gene regulator Regulator of Protease B (RopB), resulting in SIP-dependent modulation of DNA binding and regulatory activity of RopB. Notably, SIP signaling causes differential expression of ∼14% of GAS core genes. Several genes that encode toxins and other virulence genes that enhance pathogen dissemination and infection are significantly up-regulated. Using three mouse infection models, we show that the SIP signaling pathway is active during infection and contributes significantly to GAS pathogenesis at multiple host anatomic sites. Together, our results delineate the molecular mechanisms involved in a previously undescribed virulence regulatory pathway of an important human pathogen and suggest new therapeutic strategies.

RNase Y-mediated regulation of the streptococcal pyrogenic exotoxin B

“…S1B). A different organization of the intergenic region has recently been proposed: three ORFs were annotated, SPy_2040 (named orf-3 ) SIP and orf-2 [26]. In accordance with this annotation, RNA sequencing reveals that SPy_2041 is not transcribed in our experimental conditions (Fig.…”

“…1B,C). Previous studies show that the RopB binding sites are located upstream of P and P1 [23,26], indicating that RopB could promote speB expression from both promoters (Fig. S1A).…”

“…In an effort to provide coherence for future studies – the annotation of the speB 5′ UTR has been inconsistent in previous works [23,26–28] –, we first determined speB transcript boundaries. We demonstrated that RNase Y and so far unidentified RNase(s) process the 5′ UTR of speB mRNA.…”

“…A 940 nt-long intergenic region separates these two genes and contains numerous features [23]. It consists of two predicted open reading frames (ORFs) of unknown function [25] and also encodes the SpeB Inducing Peptide (SIP) involved in RopB-mediated regulation of speB expression [26]. It has recently been shown that a partial deletion of the speB 5′ UTR results in a global accumulation of mRNAs at the stationary phase of growth [27].…”

“…speB harbours a long 5′ UTR where two transcriptional start sites (TSSs), named P1 and P2, were originally predicted 697 nt and 137 nt upstream of the speB start codon, respectively [28]. Despite the important regulatory role of the intergenic region in speB expression, the annotation of this region is not consistent in the literature [21,23,26–29]. …”