Cas13b Is a Type VI-B CRISPR-Associated RNA-Guided RNase Differentially Regulated by Accessory Proteins Csx27 and Csx28
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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.
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New systems such as Cas12, Cas13a, and Cas13b are in the pipeline.
“CRISPR/Cas9 is the most widely used system, but new systems such as Cas12, Cas13a, and Cas13b are in the pipeline (Smargon et al., 2017;Zaidi et al., 2017;Aman et al., 2018).”
From Genetic resources and precise gene editing for targeted improvement of barley abiotic stress tolerance (2023) · cites this paper as Smargon et al., 2017
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- Authors
- Smargon, A. A., Cox, D. B. T., Pyzocha, N. K., Zheng, K., Slaymaker, I. M., Gootenberg, J. S., Abudayyeh, O. A., Essletzbichler, P., Shmakov, S., Makarova, K. S., Koonin, E. V., & Zhang, F.
- Affiliations
- Broad Institute (US)
- Skolkovo Institute of Science and Technology (RU)
- National Institutes of Health (US)
- Corresponding author
- Zhang, Feng
- ORCID
- Venue
- Molecular Cell
- Publisher
- Elsevier BV
- Published
- 2017-02
- Volume
- 65
- Issue
- 4
- Pages
- 618-630.e7
- ISSN
- 1097-2765 (ISSN-L)
- PMC
- PMC5432119
- OpenAlex
- W2567995016
- Cited by
- 575
- References
- 40
- Citations by year
- 2026: 36 · 2025: 56 · 2024: 68 · 2023: 52 · 2022: 74 · 2021: 76 · 2020: 74 · 2019: 59 · 2018: 45 · 2017: 34
- FWCI
- 23.67
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- Locations
- Molecular Cell (publishedVersion)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc-nd)
- PMC
- Funders
- National Institutes of Health (5rm1hg006193-10)
- National Institute of Mental Health (DP1MH100706, R01MH110049)
- National Human Genome Research Institute (RM1‐HG006193)
- National Institute of General Medical Sciences (T32GM007753)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Keywords
- collateral cleavage, RNA interference, RNA secondary structure, Escherichia coli, adaptive immunity, transcript regulation
- MeSH terms
- Gene Editing, RNA, Guide, CRISPR-Cas Systems, Bacterial Proteins, Escherichia coli, Ribonucleases, RNA, Bacterial, Gene Expression Regulation, Bacterial, Computational Biology, Databases, Genetic, RNA Interference, Data Mining, CRISPR-Cas Systems, CRISPR-Associated Proteins
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