A versatile regulatory toolkit of arabinose-inducible artificial transcription factors for Enterobacteriaceae.

The Gram-negative bacteria Salmonella enterica and Escherichia coli are important model organisms, powerful prokaryotic expression platforms for biotechnological applications, and pathogenic strains constitute major public health threats. To facilitate new approaches for research and biotechnological applications, we here develop a set of arabinose-inducible artificial transcription factors (ATFs) using CRISPR/dCas9 and Arabidopsis-derived DNA-binding proteins to control gene expression in E. coli and Salmonella over a wide inducer concentration range. The transcriptional output of the different ATFs, in particular when expressed in Salmonella rewired for arabinose catabolism, varies over a wide spectrum (up to 35-fold gene activation). As a proof-of-concept, we use the developed ATFs to engineer a Salmonella two-input biosensor strain, SALSOR 0.2 (SALmonella biosenSOR 0.2), which detects and quantifies alkaloid drugs through a measurable fluorescent output. Moreover, we use plant-derived ATFs to regulate β-carotene biosynthesis in E. coli, resulting in ~2.1-fold higher β-carotene production compared to expression of the biosynthesis pathway using a strong constitutive promoter.