Quorum sensing (QS) is a system by which bacteria communicate with each other according to the density of the population. Through this system, bacteria are able to coordinate changes in gene expression and thereby alter their phenotype to better adapt to the environment (Miller et al., 2001). Our group aims to harness this system to regulate gene expression in our modified bacteria. As more bacteria gather in the hypoxic tumour core, they sense the increase in population density, and then express the gene of interest. Therefore, by placing the gene of interest under both hypoxic and QS regulation, we can precisely regulate the expression of the therapeutic drug or gene.
The system comprises of three main components: AHL, esaR and the esaR Binding Site dependent promoter (esaRBS). AHL is a quorum sensing signal compound produced by the AHL synthase esaI (Bodman, 1995). Typically. EsaR is a luxR homologue capable of binding to promoters containing an esaRBS, silencing expression of promoted genes. This repression can be reversed through the addition of AHL to the system, which binds to esaR to form a complex with a lowered binding ability.
The core component of the QS system consists of two main parts: plac-esaR and esaRBS-GFP (Figure 1). The esaRBS part was derived from known PesaR sequences (Shong and Collins, 2013), esaR from Addgene plasmid #47660. All other components were obtained from the biobricks repository (plac, GFP, BBa_K1804001). Typically, constitutive production of esaR results in a constitutive repression of the GFP gene. However, when a critical bacterial density is reached, production of the AHL QS signal derepresses GFP production, allowing its expression and thus visual identification of activity. However, due to time constraints, we were unable to fully construct this system.
Figure 1 : Plasmid map of Plasmid map of pAC-plac-esaR-esaRBS-GFP (plac-esaR/BS-GFP).
