RT Journal Article SR Electronic T1 Programmable and portable CRISPR-Cas transcriptional activation in bacteria JF bioRxiv FD Cold Spring Harbor Laboratory SP 2020.01.03.882431 DO 10.1101/2020.01.03.882431 A1 Hsing-I Ho A1 Jennifer Fang A1 Jacky Cheung A1 Harris H. Wang YR 2020 UL http://biorxiv.org/content/early/2020/05/05/2020.01.03.882431.abstract AB Programmable gene activation enables fine-tuned regulation of endogenous and synthetic gene circuits to control cellular behavior. While CRISPR-Cas-mediated gene activation have been extensively developed for eukaryotic systems, similar strategies have been difficult to implement in bacteria. Here, we present a generalizable platform for screening and selection of functional bacterial CRISPR-Cas transcription activators. Using this platform, we identified a novel CRISPR activator, dCas9-AsiA, that could activate gene expression by up to 200-fold across genomic and plasmid targets with diverse promoters after directed evolution. The evolved dCas9-AsiA can simultaneously mediate activation and repression of bacterial regulons in E. coli. We further identified hundreds of promoters with varying basal expression that could be induced by dCas9-AsiA, which provides a rich resource of genetic parts for inducible gene activation. Finally, we show that dCas9-AsiA can be ported to other bacteria of clinical and bioindustrial relevance, thus enabling bacterial CRISPRa in more application areas. This work expands the toolbox for programmable gene regulation in bacteria and provides a useful resource for future engineering of other bacterial CRISPR-based gene regulators.Competing Interest StatementThe authors have declared no competing interest.