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Review
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CRISPR-Cas: Biology, Mechanisms and Relevance

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Review

CRISPR-Cas: Biology, Mechanisms and Relevance

Frank Hille et al. Philos Trans R Soc Lond B Biol Sci.

Abstract

Prokaryotes have evolved several defence mechanisms to protect themselves from viral predators. Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) display a prokaryotic adaptive immune system that memorizes previous infections by integrating short sequences of invading genomes-termed spacers-into the CRISPR locus. The spacers interspaced with repeats are expressed as small guide CRISPR RNAs (crRNAs) that are employed by Cas proteins to target invaders sequence-specifically upon a reoccurring infection. The ability of the minimal CRISPR-Cas9 system to target DNA sequences using programmable RNAs has opened new avenues in genome editing in a broad range of cells and organisms with high potential in therapeutical applications. While numerous scientific studies have shed light on the biochemical processes behind CRISPR-Cas systems, several aspects of the immunity steps, however, still lack sufficient understanding. This review summarizes major discoveries in the CRISPR-Cas field, discusses the role of CRISPR-Cas in prokaryotic immunity and other physiological properties, and describes applications of the system as a DNA editing technology and antimicrobial agent.This article is part of the themed issue 'The new bacteriology'.

Keywords: CRISPR; Cas9; bacteriophage; genome editing.

Figures

Figure 1.
Figure 1.
Simplified model of the immunity mechanisms of class 1 and class 2 CRISPR-Cas systems. The CRISPR-Cas systems are composed of a cas operon (blue arrows) and a CRISPR array that comprises identical repeat sequences (black rectangles) that are interspersed by phage-derived spacers (coloured rectangles). Upon phage infection, a sequence of the invading DNA (protospacer) is incorporated into the CRISPR array by the Cas1–Cas2 complex. The CRISPR array is then transcribed into a long precursor CRISPR RNA (pre-crRNA), which is further processed by Cas6 in type I and III systems (processing in type I-C CRISPR-Cas systems by Cas5d). In type II CRISPR-Cas systems, crRNA maturation requires tracrRNA, RNase III and Cas9, whereas in type V-A systems Cpf1 alone is sufficient for crRNA maturation. In the interference state of type I systems, Cascade is guided by crRNA to bind the foreign DNA in a sequence-specific manner and subsequently recruits Cas3 that degrades the displaced strand through its 3′–5′ exonucleolytic activity. Type III-A and type III-B CRISPR-Cas systems employ Csm and Cmr complexes, respectively, for cleavage of DNA (red triangles) and its transcripts (black triangles). A ribonucleoprotein complex consisting of Cas9 and a tracrRNA : crRNA duplex targets and cleaves invading DNA in type II CRISPR-Cas systems. The crRNA-guided effector protein Cpf1 is responsible for target degradation in type V systems. Red triangles represent the cleavage sites of the interference machinery.
Figure 2.
Figure 2.
Applications of the CRISPR-Cas9 technology. (a) Cas9 is guided by a sgRNA to induce a double-strand DNA break at a desired genomic locus. The DNA damage can be repaired by NHEJ yielding short random insertions or deletions at the target site. Alternatively, a DNA sequence that shows partial complementarity to the target site can be inserted during HDR for precise genome editing purposes. (b) Mutations in the catalytical domains of Cas9 yield a dead variant (dCas9) that binds but does not cleave DNA. The approach with dCas9 is used for transcriptional repression by binding to the promoter region of a gene and thus blocking the access for the RNA polymerase. Similarly, dCas9 can be fused to a transcriptional repressor. Red crosses represent inhibition of transcription. (c) The fusion of dCas9 to a transcriptional activator stimulates transcription of an adjacent gene by recruiting the RNA polymerase.

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References

    1. Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P, Moineau S, Romero DA, Horvath P. 2007. CRISPR provides acquired resistance against viruses in prokaryotes. Science 315, 1709–1712. (10.1126/science.1138140) - DOI - PubMed
    1. Haft DH, Selengut J, Mongodin EF, Nelson KE. 2005. A guild of 45 CRISPR-associated (Cas) protein families and multiple CRISPR/Cas subtypes exist in prokaryotic genomes. PLoS Comput. Biol. 1, e60 (10.1371/journal.pcbi.0010060) - DOI - PMC - PubMed
    1. Makarova KS, Grishin NV, Shabalina SA, Wolf YI, Koonin EV. 2006. A putative RNA-interference-based immune system in prokaryotes: computational analysis of the predicted enzymatic machinery, functional analogies with eukaryotic RNAi, and hypothetical mechanisms of action. Biol. Direct. 1, 7 (10.1186/1745-6150-1-7) - DOI - PMC - PubMed
    1. Mojica FJ, Diez-Villasenor C, Garcia-Martinez J, Soria E. 2005. Intervening sequences of regularly spaced prokaryotic repeats derive from foreign genetic elements. J. Mol. Evol. 60, 174–182. (10.1007/s00239-004-0046-3) - DOI - PubMed
    1. Makarova KS, et al. 2015. An updated evolutionary classification of CRISPR–Cas systems. Nat. Rev. Microbiol. 13, 722–736. (10.1038/nrmicro3569) - DOI - PMC - PubMed

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