A Type I-F Anti-CRISPR Protein Inhibits the CRISPR-Cas Surveillance Complex by ADP-Ribosylation

CRISPR-Cas systems are bacterial anti-viral systems, and phages use anti-CRISPR proteins (Acrs) to inactivate these systems. Here, we report a novel mechanism by which AcrIF11 inhibits the type I-F CRISPR system. Our structural and biochemical studies demonstrate that AcrIF11 functions as a novel mono-ADP-ribosyltransferase (mART) to modify N250 of the Cas8f subunit, a residue required for recognition of the protospacer-adjacent motif, within the crRNA-guided surveillance (Csy) complex from Pseudomonas aeruginosa. The AcrIF11-mediated ADP-ribosylation of the Csy complex results in complete loss of its double-stranded DNA (dsDNA) binding activity. Biochemical studies show that AcrIF11 requires, besides Cas8f, the Cas7.6f subunit for binding to and modifying the Csy complex. Our study not only reveals an unprecedented mechanism of type I CRISPR-Cas inhibition and the evolutionary arms race between phages and bacteria but also suggests an approach for designing highly potent regulatory tools in the future applications of type I CRISPR-Cas systems. [Display omitted] •Crystal structure of AcrIF11 suggests that it resembles an ADP-ribosyltransferase•AcrIF11 specifically ADP-ribosylates a key residue in the PAM-recognition loop•AcrIF11-mediated ADP-ribosylation of the Csy complex prevents dsDNA binding•AcrIF11 requires the Cas7.6f subunit for binding to and modifying the Csy complex Niu et al. report structural and biochemical data that reveal the molecular mechanism of AcrIF11-mediated inhibition of the type I-F CRISPR system. AcrIF11 specifically ADP-ribosylates a key residue in the PAM-recognition loop of the type I-F Cascade complex, thereby inhibiting its DNA binding activity and inactivating the CRISPR system.