Reversible Gene Expression Control in Yersinia pestis Using an Optimized CRISPRi System.

Reversible Gene Expression Control in Yersinia pestis Using an Optimized CRISPRi System. Appl Environ Microbiol. 2019 Apr 12;: Authors: Wang T, Wang M, Zhang Q, Cao S, Li X, Qi Z, Tan Y, You Y, Bi Y, Song Y, Yang R, Du Z Abstract Many genes in the bacterial pathogen Yersinia pestis, the causative agent of three plague pandemics, remain uncharacterized, greatly hampering the development of measures for plague prevention and control. Clustered regularly interspaced short palindromic repeats interference (CRISPRi) has been shown to be an effective tool for gene knockdown in model bacteria. In this system, a catalytically-dead Cas9 (dCas9) and a small guide RNA (sgRNA) form a complex, binding to the specific DNA target through base pairing, thereby impeding RNA polymerase binding and causing target gene repression. Here, we introduce an optimized CRISPRi system using Streptococcus pyogenes Cas9-derived dCas9 for gene knockdown in Y. pestis Multiple genes encoded on either the chromosome or plasmids of Y. pestis were efficiently knocked down (up to 380-fold) in a strictly anhydrotetracycline-inducible manner using this CRISPRi approach. Knockdown of hmsH (responsible for biofilm formation) or cspB (encoding a cold shock protein) resulted in greatly decreased biofilm formation or impaired cold-tolerance in in vitro phenotypic assays. Further, silencing of virulence-associated genes yscB or ail using this CRISPRi system resulted in attenuat...
Source: Applied and Environmental Microbiology - Category: Microbiology Authors: Tags: Appl Environ Microbiol Source Type: research