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TitleStructural insights into RNA-guided RNA editing by the Cas13b-ADAR2 complex.
Journal, issue, pagesNat Struct Mol Biol, Vol. 32, Issue 8, Page 1567-1578, Year 2025
Publish dateApr 11, 2025
AuthorsJunichiro Ishikawa / Kazuki Kato / Soumya Kannan / Sae Okazaki / Soh Ishiguro / Keitaro Yamashita / Nozomu Yachie / Tomohiro Nishizawa / Feng Zhang / Hiroshi Nishimasu /
PubMed AbstractCas13 is an RNA-guided RNA endonuclease derived from the type VI CRISPR-Cas system, which has been used in numerous RNA-targeting technologies, such as RNA knockdown, detection and editing. The ...Cas13 is an RNA-guided RNA endonuclease derived from the type VI CRISPR-Cas system, which has been used in numerous RNA-targeting technologies, such as RNA knockdown, detection and editing. The catalytically inactive Prevotella sp. Cas13b (dPspCas13b) fused to the human adenosine deaminase acting on RNA 2 (ADAR2) deaminase domain can edit adenosine in target transcripts to inosine, in an RNA-editing technology called REPAIR (RNA editing for programmable A-to-I replacement), which has potential for gene therapy. Here we report the cryo-electron microscopy structures of the PspCas13b-guide RNA binary complex, the PspCas13b-guide RNA-target RNA ternary complex and the dPspCas13b-ADAR2-guide RNA-target RNA complex. These structures provide mechanistic insights into RNA cleavage and editing. We applied our structural insights to engineer a compact and efficient dPspCas13b-ADAR2 complex (REPAIR-mini). Overall, our findings advance the understanding of CRISPR-Cas13 effector nucleases and could enable the development of improved RNA-targeting technologies.
External linksNat Struct Mol Biol / PubMed:40217120
MethodsEM (single particle)
Resolution2.59 - 3.71 Å
Structure data

37486

8wf8

EMDB-37486, PDB-8wf8:
Cryo-EM structure of the PspCas13b-crRNA complex
Method: EM (single particle) / Resolution: 2.59 Å

37487

8wf9

EMDB-37487, PDB-8wf9:
Cryo-EM structure of the PspCas13b-crRNA-target RNA complex (State 1)
Method: EM (single particle) / Resolution: 3.13 Å

37488

8wfa

EMDB-37488, PDB-8wfa:
Cryo-EM structure of the PspCas13b-crRNA-target RNA complex (State 2)
Method: EM (single particle) / Resolution: 3.13 Å

37489

8wfb

EMDB-37489, PDB-8wfb:
Cryo-EM structure of the dPspCas13b-ADAR2-crRNA-target RNA complex
Method: EM (single particle) / Resolution: 3.71 Å

Chemicals

ChemComp-MG:
Unknown entry

ChemComp-IHP:
INOSITOL HEXAKISPHOSPHATE

ChemComp-ZN:
Unknown entry

Source
  • prevotella sp. (bacteria)
  • synthetic construct (others)
KeywordsRNA BINDING PROTEIN/RNA / CRISPR / RNase / RNA BINDING PROTEIN-RNA COMPLEX

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