Structural basis of DNA targeting by a transposon-encoded CRISPR-Cas system

Nature. 2020 Jan;577(7789):271-274. doi: 10.1038/s41586-019-1849-0. Epub 2019 Dec 18.


Bacteria use adaptive immune systems encoded by CRISPR and Cas genes to maintain genomic integrity when challenged by pathogens and mobile genetic elements1-3. Type I CRISPR-Cas systems typically target foreign DNA for degradation via joint action of the ribonucleoprotein complex Cascade and the helicase-nuclease Cas34,5, but nuclease-deficient type I systems lacking Cas3 have been repurposed for RNA-guided transposition by bacterial Tn7-like transposons6,7. How CRISPR- and transposon-associated machineries collaborate during DNA targeting and insertion remains unknown. Here we describe structures of a TniQ-Cascade complex encoded by the Vibrio cholerae Tn6677 transposon using cryo-electron microscopy, revealing the mechanistic basis of this functional coupling. The cryo-electron microscopy maps enabled de novo modelling and refinement of the transposition protein TniQ, which binds to the Cascade complex as a dimer in a head-to-tail configuration, at the interface formed by Cas6 and Cas7 near the 3' end of the CRISPR RNA (crRNA). The natural Cas8-Cas5 fusion protein binds the 5' crRNA handle and contacts the TniQ dimer via a flexible insertion domain. A target DNA-bound structure reveals critical interactions necessary for protospacer-adjacent motif recognition and R-loop formation. This work lays the foundation for a structural understanding of how DNA targeting by TniQ-Cascade leads to downstream recruitment of additional transposase proteins, and will guide protein engineering efforts to leverage this system for programmable DNA insertions in genome-engineering applications.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Cas Systems*
  • Cryoelectron Microscopy
  • DNA Transposable Elements*
  • DNA, Bacterial / chemistry*
  • DNA, Bacterial / genetics
  • Models, Molecular
  • Nucleic Acid Conformation
  • Protein Multimerization
  • Protein Structure, Quaternary
  • RNA, Bacterial / chemistry
  • Vibrio cholerae / chemistry*
  • Vibrio cholerae / genetics


  • DNA Transposable Elements
  • DNA, Bacterial
  • RNA, Bacterial