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

Area of Science:

• Metabolic Engineering
• Synthetic Biology
• Biotechnology

Background:

• Metabolic engineering strategies often impose significant metabolic burden on host cells.
• Growth-limiting genetic modifications are most effective when activated post-growth phase.
• Controlling gene expression dynamically is crucial for optimizing cellular resource allocation.

Purpose of the Study:

• To engineer a dynamic regulatory system in Saccharomyces cerevisiae for controlled gene expression.
• To link a synthetic quorum sensing circuit with RNA interference for inducible gene silencing.
• To demonstrate the system's utility in enhancing the production of para-hydroxybenzoic acid (PHBA).

Main Methods:

• Engineered a synthetic quorum sensing circuit in Saccharomyces cerevisiae.
• Integrated an RNA interference (RNAi) module for target gene silencing.
• Utilized elementary flux mode analysis to identify optimal gene knock-downs.
• Applied dynamic RNA repression to control flux through the shikimate pathway.

Main Results:

• Successfully implemented dynamic gene silencing based on population density.
• Achieved the highest reported yeast-based para-hydroxybenzoic acid (PHBA) titer of 1.1mM.
• Enabled the application of highly productive but biomass-limiting gene modifications post-growth.

Conclusions:

• Dynamic regulation using synthetic quorum sensing and RNAi is a viable strategy in metabolic engineering.
• This approach allows for the temporal control of gene expression, optimizing production of valuable compounds.
• The developed system enhances microbial production by decoupling growth from the expression of burden-some genes.