Strategies for controlled bacterial assembly resulting in activation of a quorum-sensing circuit

Abstract

Bacterial communities show promise in a wide range of applications, such as environmental remediation and multi-step biochem. synthesis. Here we present methods by which bacteria can be assembled in two distinct architectures through controlled surface display of assocg. proteins (SpyTag and SpyCatcher or proteins contg. coiled-coil peptides including SynZip 17 and SynZip 18). We can tune the size of the aggregates by addn. of a "stoichiometric excess" of one type of cell in an interacting pair, as well as by modulating the amt. of protein displayed on the surface. We further demonstrate reversibility of the SynZip-mediated interactions, permanence of the SpyTag-SpyCatcher mediated clustering, and construction of an orthogonal assembly system using two different promoter families. We have used these systems to make simple aggregates as well as more complex core-shell structures, and we trigger quorum sensing behavior by cells within these aggregates at low global cell densities. In sum, the techniques we have developed provide a new level of control of genetically-mediated bacterial clustering, and suggest potential applications in the construction of complex bioreactor systems.