Transposon-encoded CRISPR-Cas systems direct RNA-guided DNA integration

Nature. 2019 Jul;571(7764):219-225. doi: 10.1038/s41586-019-1323-z. Epub 2019 Jun 12.

Abstract

Conventional CRISPR-Cas systems maintain genomic integrity by leveraging guide RNAs for the nuclease-dependent degradation of mobile genetic elements, including plasmids and viruses. Here we describe a notable inversion of this paradigm, in which bacterial Tn7-like transposons have co-opted nuclease-deficient CRISPR-Cas systems to catalyse RNA-guided integration of mobile genetic elements into the genome. Programmable transposition of Vibrio cholerae Tn6677 in Escherichia coli requires CRISPR- and transposon-associated molecular machineries, including a co-complex between the DNA-targeting complex Cascade and the transposition protein TniQ. Integration of donor DNA occurs in one of two possible orientations at a fixed distance downstream of target DNA sequences, and can accommodate variable length genetic payloads. Deep-sequencing experiments reveal highly specific, genome-wide DNA insertion across dozens of unique target sites. This discovery of a fully programmable, RNA-guided integrase lays the foundation for genomic manipulations that obviate the requirements for double-strand breaks and homology-directed repair.

MeSH terms

  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Base Sequence
  • CRISPR-Associated Proteins / genetics
  • CRISPR-Associated Proteins / metabolism
  • CRISPR-Cas Systems / genetics*
  • DNA Transposable Elements / genetics*
  • DNA, Bacterial / genetics*
  • DNA, Bacterial / metabolism*
  • Endodeoxyribonucleases / genetics
  • Endodeoxyribonucleases / metabolism
  • Escherichia coli / genetics
  • Gene Editing / methods*
  • Genome, Bacterial / genetics
  • Integrases / genetics
  • Integrases / metabolism
  • Mutagenesis, Insertional / methods*
  • Mutagenesis, Site-Directed / methods
  • RNA, Bacterial / genetics*
  • RNA, Guide, CRISPR-Cas Systems / genetics
  • Substrate Specificity
  • Vibrio cholerae / genetics

Substances

  • Bacterial Proteins
  • CRISPR-Associated Proteins
  • DNA Transposable Elements
  • DNA, Bacterial
  • RNA, Bacterial
  • RNA, Guide, CRISPR-Cas Systems
  • Integrases
  • Endodeoxyribonucleases