title: Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression.
abstract: CRISPR-Cas systems have shown tremendous promise as heterologous tools for genome editing and transcriptional regulation. Because these RNA-directed immune systems are found in most prokaryotes, an opportunity exists to harness the endogenous systems as convenient tools in these organisms. Here, we report that the Type I-E CRISPR-Cas system in Escherichia coli can be co-opted for programmable transcriptional repression. We found that deletion of the signature cas3 gene converted this immune system into a programmable gene regulator capable of reversible gene silencing of heterologous and endogenous genes. Targeting promoter regions yielded the strongest repression, whereas targeting coding regions showed consistent strand bias. Furthermore, multi-targeting CRISPR arrays could generate complex phenotypes. This strategy offers a simple approach to convert many endogenous Type I systems into transcriptional regulators, thereby expanding the available toolkit for CRISPR-mediated genetic control while creating new opportunities for genome-wide screens and pathway engineering.
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC 27695, USA.
authors.forename: Michelle L
authors.initials: ML
authors.lastname: Luo
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC 27695, USA.
authors.forename: Adam S
authors.initials: AS
authors.lastname: Mullis
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC 27695, USA.
authors.forename: Ryan T
authors.initials: RT
authors.lastname: Leenay
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC 27695, USA cbeisel@ncsu.edu.
authors.forename: Chase L
authors.initials: CL
authors.lastname: Beisel
mesh_terms: D064130:CRISPR-Associated Proteins / Q000235:genetics; D064113:CRISPR-Cas Systems; D060846:Cell Engineering; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D014158:Transcription, Genetic
chemical_list: D064130:CRISPR-Associated Proteins