Home
This Title All WIREs
WIREs RSS Feed
How to cite this WIREs title:
WIREs RNA
Impact Factor: 5.844

CRISPR‐Cas systems and RNA‐guided interference

Full article on Wiley Online Library:   HTML PDF

Can't access this content? Tell your librarian.

Clustered regularly interspaced short palindromic repeats (CRISPR) together with associated sequences (cas) form the CRISPR‐Cas system, which provides adaptive immunity against viruses and plasmids in bacteria and archaea. Immunity is built through acquisition of short stretches of invasive nucleic acids into CRISPR loci as ‘spacers'. These immune markers are transcribed and processed into small noncoding interfering CRISPR RNAs (crRNAs) that guide Cas proteins toward target nucleic acids for specific cleavage of homologous sequences. Mechanistically, CRISPR‐Cas systems function in three distinct stages, namely: (1) adaptation, where new spacers are acquired from invasive elements for immunization; (2) crRNA biogenesis, where CRISPR loci are transcribed and processed into small interfering crRNAs; and (3) interference, where crRNAs guide the Cas machinery to specifically cleave homologous invasive nucleic acids. A number of studies have shown that CRISPR‐mediated immunity can readily increase the breadth and depth of virus resistance in bacteria and archaea. CRISPR interference can also target plasmid sequences and provide a barrier against the uptake of undesirable mobile genetic elements. These inheritable hypervariable loci provide phylogenetic information that can be insightful for typing purposes, epidemiological studies, and ecological surveys of natural habitats and environmental samples. More recently, the ability to reprogram CRISPR‐directed endonuclease activity using customizable small noncoding interfering RNAs has set the stage for novel genome editing and engineering avenues. This review highlights recent studies that revealed the molecular basis of CRISPR‐mediated immunity, and discusses applications of crRNA‐guided interference. WIREs RNA 2013, 4:267–278. doi: 10.1002/wrna.1159

This article is categorized under:

  • RNA Turnover and Surveillance > Turnover/Surveillance Mechanisms
  • Regulatory RNAs/RNAi/Riboswitches > Biogenesis of Effector Small RNAs
  • Regulatory RNAs/RNAi/Riboswitches > RNAi: Mechanisms of Action

This WIREs title offers downloadable PowerPoint presentations of figures for non-profit, educational use, provided the content is not modified and full credit is given to the author and publication.

Download a PowerPoint presentation of all images


Figure 1.

CRISPR‐Cas elements and mechanism of action. The various elements that constitute CRISPR‐Cas systems are graphically depicted, including cas genes (with the universal cas1 and cas2), the leader (L), spacers (boxes), repeats (diamonds). Adaptive immunity is build through acquisition of new spacers at the leader end, which are derived from invasive DNA (here, phage DNA). In the expression stage, the repeat‐spacer array is then transcribed as a full‐length pre‐crRNA, which is processed by cleavage into mature crRNAs. The mature RNAs then form a ribonucleoprotein complex with Cas proteins to mediate interference by guiding endonucleases toward homologous nucleic acid sequences.

[ Normal View | Magnified View ]
Figure 2.

Type I CRISPR‐Cas systems. Details are provided for the prototypical Escherichia coli K12 type I‐E system (ygcB‐ygbF). The universal cas1 and cas2 genes are shown in black. The type I signature gene, cas3, is shown in red, while the crRNA maturation nuclease cas6 is shown in green. Repeats are represented as black diamonds, and spacers as white boxes.

[ Normal View | Magnified View ]
Figure 3.

Type II CRISPR‐Cas systems. Details are provided for the prototypical Streptococcus thermophilus DGCC7710 type II‐A system. The universal cas1 and cas2 genes are shown in black. The type II signature gene, cas9, is shown in purple. Repeats are represented as black diamonds, and spacers as white boxes.

[ Normal View | Magnified View ]
Figure 4.

Type III CRISPR‐Cas systems. Details are provided for the prototypical Pyrococcus furiosus type III‐B system. The universal cas1 and cas2 genes are shown in black. The type III signature gene, cas10, is shown in orange, while the crRNA maturation nuclease cas6 is shown in green. Repeats are represented as black diamonds, and spacers as white boxes.

[ Normal View | Magnified View ]
Figure 5.

crRNA‐mediated DNA cleavage. (a) Sequences and details are provided for the Streptococcus thermophilus CRISPR3‐Cas system, which contains 36nt CRISPR repeats and 30nt spacers. crRNA biogenesis requires Cas9, RNaseIII and tracrRNA. (b) dsDNA cleavage is driven by RuvC and HNH strand nicking within a R‐loop, 3 nt away from the 3′ terminus of the protospacer, which is flanked by the 5′‐NGGNG‐3′ proto‐spacer adjacent motif (PAM). Rather than used the natural Cas9‐crRNA‐tracrRNA system, it is possible to generate a chimeric RNA which mimics the crRNA‐tracrRNA duplex for a two‐component flexible system. (c) Because RuvC and HNH each nick one strand of the target DNA, it is possible to reprogram the Cas9 endonuclease to either generate a dsDNA break, or a nick on either strand exactly 3 bp upstream of the PAM. Using the WT Cas9 endonuclease, dsDNA cleavage is generated (top). Using a HNH‐ mutant, the (−) strand is cleaved (bottom left), whereas using a RuvC‐ mutant, the (+) strand is nicked (bottom right). Likewise, any crRNA targeting a sequence flanked by the PAM can be used to reprogram the Cas9 nuclease.

[ Normal View | Magnified View ]

Related Articles

Regulatory Non-Coding RNAs

Browse by Topic

Regulatory RNAs/RNAi/Riboswitches > Biogenesis of Effector Small RNAs
Regulatory RNAs/RNAi/Riboswitches > RNAi: Mechanisms of Action
RNA Turnover and Surveillance > Turnover/Surveillance Mechanisms

Access to this WIREs title is by subscription only.

Recommend to Your
Librarian Now!

The latest WIREs articles in your inbox

Sign Up for Article Alerts