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C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector.

Omar O Abudayyeh1, Jonathan S Gootenberg2, Silvana Konermann3

  • 1Department of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. McGovern Institute for Brain Research at MIT, Cambridge, MA 02139, USA. Departments of Brain and Cognitive Science and Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.

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Researchers characterized the C2c2 CRISPR-Cas effector, a novel RNA-guided ribonuclease. This system from Leptotrichia shahii targets and cleaves specific RNA molecules, offering new RNA-targeting tools.

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Biochemistry

Background:

  • The clustered regularly interspaced short palindromic repeat (CRISPR)-CRISPR-associated genes (Cas) system provides adaptive immunity in microbes.
  • CRISPR-Cas systems function through DNA or RNA-DNA interference against foreign genetic elements.
  • Class 2 CRISPR-Cas effectors represent a distinct category of these microbial defense mechanisms.

Purpose of the Study:

  • To characterize the class 2 type VI CRISPR-Cas effector, C2c2.
  • To demonstrate the RNA-guided ribonuclease activity of C2c2.
  • To explore the potential of C2c2 as a novel RNA-targeting tool.

Main Methods:

  • Biochemical analysis of C2c2 from Leptotrichia shahii.
  • In vitro assays to assess RNA cleavage activity.
  • Mutagenesis of conserved Higher Eukaryotes and Prokaryotes Nucleotide-binding (HEPN) domains.

Main Results:

  • C2c2 exhibits RNA-guided ribonuclease activity, targeting single-stranded RNA.
  • C2c2 mediates interference against RNA phage in bacteria.
  • Specific mRNA knockdown was achieved in bacteria by programming C2c2.
  • Catalytic activity is dependent on conserved residues within the HEPN domains.

Conclusions:

  • C2c2 is a versatile RNA-guided ribonuclease with potential applications in RNA targeting.
  • Understanding C2c2 expands the knowledge of CRISPR-Cas system diversity and function.
  • C2c2 can be engineered into novel tools for RNA manipulation.