Stably Integrating an Inducible CRISPR-Cas9 to Protect Against Viral Infections in Vitro.

ABSTRACT

CRISPR-Cas systems protect bacteria from viral nucleic acids. The Cas9 enzyme cleaves bacteriophage DNA preventing viral genes from being expressed in the bacterial host. In this work, the Cas9 protein is repurposed to function as an intracellular mammalian defense mechanism that protects human cells from cytomegaloviral DNA. The A549 lung adenocarcinoma cell line was genetically modified to express a doxycycline-inducible Cas9, and a guide RNA targeting a luciferase reporter plasmid. This investigation revealed a robust inducible Cas9 system that successfully reduced the expression of the luciferase viral reporter by up to 98% and by 75% on average.