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TitleExploiting activation and inactivation mechanisms in type I-C CRISPR-Cas3 for genome-editing applications.
Journal, issue, pagesMol Cell, Vol. 84, Issue 3, Page 463-475.e5, Year 2024
Publish dateFeb 1, 2024
AuthorsChunyi Hu / Mason T Myers / Xufei Zhou / Zhonggang Hou / Macy L Lozen / Ki Hyun Nam / Yan Zhang / Ailong Ke /
PubMed AbstractType I CRISPR-Cas systems utilize the RNA-guided Cascade complex to identify matching DNA targets and the nuclease-helicase Cas3 to degrade them. Among the seven subtypes, type I-C is compact in size ...Type I CRISPR-Cas systems utilize the RNA-guided Cascade complex to identify matching DNA targets and the nuclease-helicase Cas3 to degrade them. Among the seven subtypes, type I-C is compact in size and highly active in creating large-sized genome deletions in human cells. Here, we use four cryoelectron microscopy snapshots to define its RNA-guided DNA binding and cleavage mechanisms in high resolution. The non-target DNA strand (NTS) is accommodated by I-C Cascade in a continuous binding groove along the juxtaposed Cas11 subunits. Binding of Cas3 further traps a flexible bulge in NTS, enabling NTS nicking. We identified two anti-CRISPR proteins AcrIC8 and AcrIC9 that strongly inhibit Neisseria lactamica I-C function. Structural analysis showed that AcrIC8 inhibits PAM recognition through allosteric inhibition, whereas AcrIC9 achieves so through direct competition. Both Acrs potently inhibit I-C-mediated genome editing and transcriptional modulation in human cells, providing the first off-switches for type I CRISPR eukaryotic genome engineering.
External linksMol Cell / PubMed:38242128 / PubMed Central
MethodsEM (single particle)
Resolution3.0 - 3.64 Å
Structure data

29877

8g9s

EMDB-29877, PDB-8g9s:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.4 Å

29878

8g9t

EMDB-29878, PDB-8g9t:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.6 Å

29879

8g9u

EMDB-29879, PDB-8g9u:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.0 Å

29896

8gaf

EMDB-29896, PDB-8gaf:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.64 Å

29900

8gam

EMDB-29900, PDB-8gam:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.46 Å

29901

8gan

EMDB-29901, PDB-8gan:
Exploiting Activation and Inactivation Mechanisms in Type I-C CRISPR-Cas3 for Genome Editing Applications
Method: EM (single particle) / Resolution: 3.26 Å

Chemicals

ChemComp-MG:
Unknown entry

ChemComp-PO4:
PHOSPHATE ION / Phosphate

Source
  • neisseria lactamica (bacteria)
  • rhodobacter phage rcnl1 (virus)
KeywordsHYDROLASE/RNA / CRISPR / type I-C / cascade / anti-CRISPR / HYDROLASE-RNA complex / DNA BINDING PROTEIN / HYDROLASE/RNA/DNA / HYDROLASE-RNA-DNA complex

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