The miniature CRISPR-Cas12m effector binds DNA to block transcription

Wen Y. Wu*, Prarthana Mohanraju, Chunyu Liao, Belén Adiego-Pérez, Sjoerd C.A. Creutzburg, Kira S. Makarova, Karlijn Keessen, Timon A. Lindeboom, Tahseen S. Khan, Stijn Prinsen, Rob Joosten, Winston X. Yan, Anzhela Migur, Charlie Laffeber, David A. Scott, Joyce H.G. Lebbink, Eugene V. Koonin, Chase L. Beisel, John van der Oost

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

27 Citations (Scopus)

Abstract

CRISPR-Cas are prokaryotic adaptive immune systems. Cas nucleases generally use CRISPR-derived RNA guides to specifically bind and cleave DNA or RNA targets. Here, we describe the experimental characterization of a bacterial CRISPR effector protein Cas12m representing subtype V-M. Despite being less than half the size of Cas12a, Cas12m catalyzes auto-processing of a crRNA guide, recognizes a 5′-TTN′ protospacer-adjacent motif (PAM), and stably binds a guide-complementary double-stranded DNA (dsDNA). Cas12m has a RuvC domain with a non-canonical catalytic site and accordingly is incapable of guide-dependent cleavage of target nucleic acids. Despite lacking target cleavage activity, the high binding affinity of Cas12m to dsDNA targets allows for interference as demonstrated by its ability to protect bacteria against invading plasmids through silencing invader transcription and/or replication. Based on these molecular features, we repurposed Cas12m by fusing it to a cytidine deaminase that resulted in base editing within a distinct window.

Original languageEnglish
Pages (from-to)4487-4502.e7
JournalMolecular Cell
Volume82
Issue number23
Early online date24 Nov 2022
DOIs
Publication statusPublished - 1 Dec 2022

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© 2022 Elsevier Inc.

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