TY - JOUR T1 - Mechanism of broad-spectrum Cas9 inhibition by AcrIIA11 JF - bioRxiv DO - 10.1101/2021.09.15.460536 SP - 2021.09.15.460536 AU - Kaylee E. Dillard AU - Cynthia Terrace AU - Kamyab Javanmardi AU - Wantae Kim AU - Kevin J. Forsberg AU - Ilya J. Finkelstein Y1 - 2021/01/01 UR - http://biorxiv.org/content/early/2021/09/15/2021.09.15.460536.abstract N2 - Mobile genetic elements evade CRISPR-Cas adaptive immunity by encoding anti-CRISPR proteins (Acrs). Acrs inactivate CRISPR-Cas systems via diverse mechanisms but are generally specific for a narrow subset of Cas nucleases that share high sequence similarity. Here, we demonstrate that AcrIIA11 inhibits diverse Cas9 sub-types in vitro and human cells. Single-molecule fluorescence imaging reveals that AcrIIA11 interferes with the first steps of target search by reducing S. aureus Cas9’s diffusion on non-specific DNA. DNA cleavage is inhibited because the AcrIIA11:Cas9 complex is kinetically trapped at PAM-rich decoy sites, preventing Cas9 from reaching its target. This work establishes that DNA trapping can be used to inhibit a broad spectrum of Cas9 orthologs in vitro and during mammalian genome editing.Competing Interest StatementThe authors have declared no competing interest. ER -