[YSA] Young Scientist Award Symposium

[YSA-7(2P-20)] Structure of the miniature type V-F CRISPR-Cas effector enzyme

^○Satoru Takeda (Dept. of Biol. Sci., Grad. Sch. of Sci., Univ. of Tokyo)

RNA-guided DNA endonucleases derived from CRISPR-Cas adaptive immune systems are widely used as powerful genome-engineering tools. Among the diverse CRISPR-Cas nucleases, the type V-F Cas12f proteins are exceptionally compact, and associate with a guide RNA to cleave single- and double-stranded DNA targets. Here, we report the cryo-electron microscopy structure of Cas12f1 (hereafter referred to as Cas12f for simplicity) in complex with a guide RNA and its target DNA. Unexpectedly, the structure revealed that two Cas12f molecules assemble with the single guide RNA to recognize the double-stranded DNA target. Each Cas12f protomer adopts a different conformation and plays distinct roles in the nucleic-acid recognition and DNA cleavage, thereby explaining how the miniature Cas12f enzyme achieves RNA-guided DNA cleavage as an "asymmetric homodimer". Our findings augment the mechanistic understanding of diverse CRISPR-Cas nucleases, and provide a framework for the development of compact genome-engineering tools critical for therapeutic genome editing.