Cas13b Is a Type VI-B CRISPR-Associated RNA-Guided RNase Differentially Regulated by Accessory Proteins Csx27 and Csx28.

Mol Cell
Authors
Abstract

CRISPR-Cas adaptive immune systems defend microbes against foreign nucleic acids via RNA-guided endonucleases. Using a computational sequence database mining approach, we identify two class 2 CRISPR-Cas systems (subtype VI-B) that lack Cas1 and Cas2 and encompass a single large effector protein, Cas13b, along with one of two previously uncharacterized associated proteins, Csx27 and Csx28. We establish that these CRISPR-Cas systems can achieve RNA interference when heterologously expressed. Through a combination of biochemical and genetic experiments, we show that Cas13b processes its own CRISPR array with short and long direct repeats, cleaves target RNA, and exhibits collateral RNase activity. Using an E. coli essential gene screen, we demonstrate that Cas13b has a double-sided protospacer-flanking sequence and elucidate RNA secondary structure requirements for targeting. We also find that Csx27 represses, whereas Csx28 enhances, Cas13b-mediated RNA interference. Characterization of these CRISPR systems creates opportunities to develop tools to manipulate and monitor cellular transcripts.

Year of Publication
2017
Journal
Mol Cell
Volume
65
Issue
4
Pages
618-630.e7
Date Published
2017 Feb 16
ISSN
1097-4164
DOI
10.1016/j.molcel.2016.12.023
PubMed ID
28065598
Links
Grant list
DP1 MH100706 / MH / NIMH NIH HHS / United States
R01 MH110049 / MH / NIMH NIH HHS / United States
RM1 HG006193 / HG / NHGRI NIH HHS / United States