Engineered Bacillus subtilis Systems for the Characterization and Detection of Peptide-Based Quorum Sensing in...

Many questions remain about the biochemical mechanisms of quorum sensing (QS), particularly in Gram-positive bacterial pathogens that pose serious threats to human health such as Staphylococcus aureus. Methods to isolate and study the molecular components of these signaling systems in vitro are not straightforward. To date, most mechanistic investigations into QS systems have been accomplished in cells and, to a lesser extent, using chemical inhibitors. Herein, we report the development and characterization of a set of engineered Bacillus subtilis strains that can be used to study the accessory gene regulator (agr) QS circuits of a variety of Gram-positive bacteria. We cloned the S. aureus agr QS machinery into B. subtilis as proof-of-concept to generate "sender" cells capable of autoinducing peptide (AIP) signal biosynthesis, "receiver" cells capable of signal transduction, and "full" cells containing the full agr QS system. We verified that the B. subtilis sender cells could produce the native S. aureus AIP and that the receiver and full cells responded to both the AIP signal and a panel of known chemical agr modulators. The approach was readily transferable to study the agr QS systems of Staphylococcus epidermidis and Listeria monocytogenes, revealing interesting differences that could indicate underlying divergences in native QS mechanisms. We also demonstrate the ability of these B. subtilis strains to function as biosensors to detect the native AIPs of bacteria and produce a targeted antibiotic in response. These engineered systems should find utility for the study of QS in a range of fundamental and applied contexts.