Inhibition of CRISPR Cas9 with Bacteriophage Proteins
Bacterial CRISPR Cas systems utilize sequence specific RNA guided nucleases to defend against bacteriophage infection. As a countermeasure, numerous phages are known that produce proteins to block the function of class 1 CRISPR Cas systems. However, currently no proteins are known to inhibit the widely used class 2 CRISPR Cas9 system. To find these inhibitors, we searched cas9 containing bacterial genomes for the co existence of a CRISPR spacer and its target, a potential indicator for CRISPR inhibition. This analysis led to the discovery of four unique type II A CRISPR Cas9 inhibitor proteins encoded by Listeria monocytogenes prophages. More than half of L. monocytogenes strains with cas9 contain at least one prophage encoded inhibitor, suggesting widespread CRISPR Cas9 inactivation. Two of these inhibitors also blocked the widely used Streptococcus pyogenes Cas9 when assayed in Escherichia coli and human cells. These natural Cas9 specific "anti CRISPRs" present tools that can be used to regulate the genome engineering activities of CRISPR Cas9.
Cell