CRISPR-Cas Activation (CRISPRa) for Transcriptional Control in E. coli

Metabolic engineering efforts often require controlling the expression level of multiple genes. We have developed a new suite of synthetic bacterial transcriptional activators by linking activation domains to the CRISPR-Cas system. Together with existing tools for CRISPRi gene repression, these synthetic activators enable programmable transcriptional up- and down-regulation over multiple endogenous and heterologous targets at the same time. Interestingly, effective gene activation requires target sites situated in a narrow region just upstream of the gene of interest, in sharp contrast to the relatively flexible target site requirements for gene activation in eukaryotic cells. We present a strategy for synthetic promoter design to independently activate multiple heterologous genes via CRISPRa. Last, we demonstrate the bioproduction of industrially-relevant aromatics (p-aminocinnamic acid) via CRISPRa control of heterologous gene expression in E. coli. In this talk, I will present these results and describe on-going work to fine-tune CRISPRa transcriptional activation, engineer dynamically-controlled CRISPRa systems and build more complex synthetic bacterial devices with a wide range of applications in industrial biosynthesis.