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TitleRNA anti-CRISPRs deplete Cas proteins to inhibit the CRISPR-Cas system.
Journal, issue, pagesMol Cell, Year 2025
Publish dateDec 30, 2025
AuthorsXiaopan Gao / Kaixiang Zhu / Weihe Zhang / Lin Wang / Linyue Wang / Lei Hua / Tongxin Niu / Bo Qin / Xia Yu / Hongtao Zhu / Sheng Cui /
PubMed AbstractRNA-based anti-CRISPRs (Racrs) interfere with the type I-F CRISPR-Cas system by mimicking the repeats found in CRISPR arrays. Here, we determined the cryo-electron microscopy (cryo-EM) structures of ...RNA-based anti-CRISPRs (Racrs) interfere with the type I-F CRISPR-Cas system by mimicking the repeats found in CRISPR arrays. Here, we determined the cryo-electron microscopy (cryo-EM) structures of the type I-F crRNA-guided surveillance complex (Csy complex) from Pectobacterium atrosepticum and three RacrIF1-induced aberrant subcomplexes. Additionally, we observed that Cas7f proteins could bind to non-specific nucleic acids, forming right-handed superhelical filaments composed of different Cas7 copies. Mechanistically, RacrIF1 lacks the specific S-conformation observed in the corresponding position of the 5' handle in canonical CRISPR complexes, and it instead adopts a periodic "5 + 1" pattern. This conformation creates severe steric hindrance for Cas5f-Cas8f heterodimer and undermines their binding. Furthermore, Cas7f nonspecifically binds nucleic acids and can form infinite superhelical filaments along Racrs molecules. This oligomerization sequesters Cas6f and Cas7f from binding, therefore blocking the formation of functional CRISPR-Cas effector complexes and ultimately blocking antiviral immunity. Our study provides a structural basis underlying Racrs-mediated CRISPRs inhibition.
External linksMol Cell / PubMed:41475348
MethodsEM (single particle)
Resolution2.8 - 4.85 Å
Structure data

60812

9irf

EMDB-60812, PDB-9irf:
Cryo-EM Structure of csy1-4 with crRNA
Method: EM (single particle) / Resolution: 2.8 Å

60813

9irg

EMDB-60813, PDB-9irg:
Cryo-EM Structure of RNA
Method: EM (single particle) / Resolution: 4.59 Å

60815

9iri

EMDB-60815, PDB-9iri:
Cryo-EM Structure of D-RNA
Method: EM (single particle) / Resolution: 2.8 Å

EMDB-60817: Cryo-EM Structure of rRNA
Method: EM (single particle) / Resolution: 3.7 Å

EMDB-60819: Cryo-EM Structure of CRISPR
Method: EM (single particle) / Resolution: 3.2 Å

66729

9xcf

EMDB-66729, PDB-9xcf:
Cryo-EM structure of csy3 with crRNA
Method: EM (single particle) / Resolution: 4.58 Å

66731

9xcg

EMDB-66731, PDB-9xcg:
Cryo-EM structure of csy3 with crRNA
Method: EM (single particle) / Resolution: 4.85 Å

Source
  • pectobacterium atrosepticum scri1043 (bacteria)
  • thiocystis violascens (bacteria)
  • thiocystis violascens dsm 198 (bacteria)
KeywordsRNA BINDING PROTEIN/RNA / crRNA / cas6f / cas7f / cas5f / cas8f / RNA BINDING PROTEIN-RNA complex / IMMUNE SYSTEM/RNA / CRISPR-Cas system / IMMUNE SYSTEM-RNA complex

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