Quorum-sensing linked RNA interference for dynamic metabolic pathway control in Saccharomyces cerevisiae

• The most productive metabolic engineering strategies often prevent cell growth.
• Dynamic regulation enables these strategies after biomass accumulation.
• Quorum sensing linked RNAi was used to dynamically regulate PHBA production.
• Lethal gene knockouts were replaced by dynamic repression to increase PHBA titer.
• This synthetic circuit can potentially be used to control any pathway in yeast.

Some of the most productive metabolic engineering strategies involve genetic modifications that cause severe metabolic burden on the host cell. Growth-limiting genetic modifications can be more effective if they are ‘switched on’ after a population growth phase has been completed. To address this problem we have engineered dynamic regulation using a previously developed synthetic quorum sensing circuit in Saccharomyces cerevisiae. The circuit autonomously triggers gene expression at a high population density, and was linked with an RNA interference module to enable target gene silencing. As a demonstration the circuit was used to control flux through the shikimate pathway for the production of para-hydroxybenzoic acid (PHBA). Dynamic RNA repression allowed gene knock-downs which were identified by elementary flux mode analysis as highly productive but with low biomass formation to be implemented after a population growth phase, resulting in the highest published PHBA titer in yeast (1.1 mM).